Empowerment on Girl Child Essay Example for Free

Empowerment on Girl Child Essay With sacrificing family resources to educate a girl child and a potential future leader still a big societal challenge, any effort to see the education of a girl is a huge boon. So when millions of dollars are poured into the effort, the impact cannot be overemphasised. The Campaign for Female Education (Camfed), introduced some few years back, has seen remarkable change of fortunes to many a rural folk. Now, a US$19 million bursary programme has been launched to benefit 24 000 girls from disadvantaged families in rural Guruve, Mashonaland Central. The launch was conducted at colourful ceremony at Chifamba Secondary School in the area recently. With testimonies of previous beneficiaries of the programme giving the clear picture of changed lives, more girls are set to change for the better. Already, lives have changed and tales are being told. More are coming, definitely. Twenty-five-year-old Bridget Moyo was born in the dusty village of Wedza in a polygamous family. Her mother sired six children and the other children under the genealogy of her father are incalculable. She needs to sit down and count them from her father’s first wife until the last. Being a girl on a polygamous family, she was not spared from challenges women as a whole face. From birth she was automatically rendered a future beggar. Her education was considered optional and it was the first thing to be sacrificed in a crisis. Her brothers, uncles and male cousins’ needs had to come first for the family. The family’s future was seen to be in their hands and blood, so it was to them that the family’s resources should be spent primarily. As if that was not enough, the family was so much immersed in poverty. School fees and levies were a luxury they could only dream of and there wasn’t enough for the family to eat. â€Å"I lost count of how many other people’s fields we worked in to make ends meet with my mother. It was not unusual for people to approach my mother and offer me a job as their housemaid,† Bridget said. She said it was very tragic in that some people had the audacity to exchange her labour services with a bucket of maize a month. â€Å"I am a proud member of the Johane Marange Apostolic Sect and my growing up in the church came with benefits and challenges. â€Å"I feel at home hen at church where I am accepted with expectations like other girls who have to get married at a tender age. † â€Å"In my teenage years, I was only supposed to dream about the kind of husband I was going to marry. Even if it meant dropping out of school, I did not drop out until I attained my university degree,† Bridget went on to narrate her ordeal. The turning point in Bridget’s life came after she got a bursary before attending secondary education. â€Å"In primary school I vividly remember being nominated a prefect before the school authorities reversed the decision because I did not have a school uniform. I never had a worry about the strategy to use to sneak back into classroom after being sent home on numerous occasions to collect the fees . Currently I am a holder of Bachelor of Science Honours Degree in Business Management and Entrepreneurship,† she said. This is not the only sad tale about girls who rise from invisibility to visibility after attaining education. Another is Talent Tokoda, who grew up as an orphan and single child. Talent was born and bred in Chivhu, where her mother took care of all the family needs. â€Å"It was a nightmare getting shoes or having a proper uniform. I struggled through primary school to completion but fortunately I passed with five units which are a sharp contrast to the struggles I went through. † â€Å"Time to enrol for secondary education came and my hope was like a dim light at the far end of a tunnel which could turn off anytime. A week before I was supposed to go to secondary school, I neither had school fees nor secured a place at any school. † â€Å"I could spend the whole day in the garden with my mother. I got the surprise of my life when I was told that my fees were going to be paid for until I complete Advanced Level,† Talent said in front of the dumbfounded crowd. She passed Advanced Level and was enrolled at the University of Zimbabwe where she is doing her final year studying for a Bachelor in Medicine and Surgery. â€Å"I am proud that I proved to doomsayers that I can achieve any goal men can achieve. In August next year I will be a qualified medical doctor,† Talent said in front of the cheering crowd. This mirrors how the personality can be moulded to greatness. Guruve District’s pass rate is pegged at 25 percent with the national pass rate sitting at 21 percent. Assisting the girl child with resources will help improve the pass rate at rural schools. For example, at Chifamba Secondary School the pass rate for girls is pegged at 10 percent. Research revealed that in Sub-Saharan Africa, 24 million girls cannot afford to go to school and as a result a girl may marry as young as 13. Camfed executive director for Zimbabwe and Malawi, Ms Angeline Murimirwa said it is vital to improve educational access, progression and completion for marginalised secondary school girls. â€Å"The coverage of bursaries will span for four years in 28 rural districts including resettlement areas. The other money will provide a package of support to schools, training of school development committees and support for parents to enable children currently out of school to enrol,† she said. Ms Murimirwa said it is imperative to enhance participation of women in national activities from district level. â€Å"Most secondary school girls drop out of school opting to get married or as a result of lacking financial support. â€Å"Organisations need to cherish marginalised communities and the idea that women constitute a greater percentage to the national population,† she said.

Overview Of Using Enzymes In Biocatalysis Green Chemistry Biology Essay

Overview Of Using Enzymes In Biocatalysis Green Chemistry Biology Essay ABSTRACT Aminotransaminases are a group of enzymes that play a key role in the amino acid metabolism catalysing the transfer of amino groups into keto acids resulting in the production of amino derivatives. It is PLP-dependent.they have many applications as biocatalysts because of their ability to introduce amino into ketone with good enantio- and regioselectivity. Aminotransaminases have been classified by three groups on the basis of substrate specifity, PLP-fold similarity and structural similarities. One of the important sub-groups of transaminases is omega transaminase (à Ã¢â‚¬ °-AT) which is capable of transferring amino group from a primary amine that does not contain a carboxyl group. There are many available à Ã¢â‚¬ °-ATs but the first crystal structure of this type of enzyme identified from Chromobacterium violaceum. Keywords: Aminotransaminase; PLP-dependent; regioselectivity; chromobacterium violaceum. INTRODUCTION Transaminases or aminotransferases are the group of the transferase enzymes which are involved in the reversible transfer of amino groups from amino acid to ÃŽÂ ±-keto acids. The enzyme uses pyridoxal-5-phosphate (PLP) in the reaction, therefore, it has been classified under PLP-dependent enzymes (Mehta et al., 1993). In current decades the importance of transaminases have significantly increased as a result of their tremendous potential for the production of both natural and unnatural amino acids and enantiomerically pure chiral amines which are important particularly for pharmaceutical industry (Shin et al., 2000). This review will mainly focus on the structure, mechanism and biotechnological application of omega transaminase (à Ã¢â‚¬ °-AT) enzymes from different sources. The first section will give a general overview of using enzyme in white biotechnology. The second part will give general overview of transaminases with different classes of transaminases. In the following part, general reaction mechanism of transaminase and structure of the chromobacterial omega transaminase will be explained in detail. Under the last subheading biotechnological application of omega transaminase will be discussed. Finally it will give the project aims and conclusion. Overview of using enzymes in biocatalysis/green chemistry Enzymes are fundamental catalysts which are capable of acting on a wide range of complex compounds as substrates. They are exquisitely selective catalysing reaction leading to production enantio- and regio-selective intermediates (Schmid et al., 2001). Isolated enzymes and whole cell biocatalysts are commonly used to produce optically pure compounds. Isolated enzymes are generally used for the aim of the catalysis of hydrolytic and isomerisation reactions; whereas; whole cells are typically used for synthetic reactions (Schmid et al., 2001). Both of them have some advantages and disadvantages. The disadvantages of using whole cells in the biocatalysis reactions are varied. For instance, substrate molecule might be toxic and results to death of cell; or the size of substrate molecule might be so huge that cannot pass through membrane; or there may be other enzymes in the cell that acts on the same substrate and cause to the production more than one compound (Wubbolts et al., 1994). In spite of these disadvantages, no requirement for recycling process of co-enzyme makes them good candidate to be used in biocatalysis because of the economic factors. The rapid increase in the development of research area of protein engineering, including molecular evolution, and enzyme engineering, has resulted in rapid growth of biocatalysis. The protein engineering yields molecule with modified structure, function and selectivity, in aqueous environment; whereas, the enzyme engineering leads to remarkable improvement particularly in organic solvent. Using organic solvent provides many advantages such as higher substrate solubility, modified enzyme specifity that results in the new and higher enzymatic activity that previously were only feasible using genetic modifications or complex reaction pathways inside the cell. As a consequence, applications of biocatalysis in organic environment vary from chiral resolution of pharmaceutical intermediates, chemical compounds to enantio- and regioselective polymerisation (Schmid et al., 2001). The use of biocatalysis in the industry for the synthesis of synthetic compounds has been significantly increased as the use of biocatalytic process for producing industrial intermediate has become easier. Biocatalytic reactions can be performed in the organic solvents and also water. This allows selective and efficient conversion of both water soluble and apolar organic molecules using biocatalytically active cells or molecules. The production of optically active substances is an area of growing demand in the fine chemical industry and biocatalysis has developed from a niche technology to a commonly used manufacturing method. The selectivity and cushy operational conditions of biocatalysists are increasingly applied in industry to modify complex target molecules. (Panke et al., 2004). General Overview of Transaminases Different classes of transaminases -classification Aminotransferases (EC 2.6.1.X) are the group of enzymes that take a significant role in the transamination reactions. They involve in the exchange of oxygen from alfa keto acid and amine from an amino acid, thus, they remove the amino group from the amino acid and transfer it to alfa keto acid and converting it into amino acid (Mehta et al., 1993). Using aminotransferases in the biocatalysis provides many advantages over other group enzymes for the production of chiral compounds. The reasons which make them so attractive are being able to act on wide range of substrate, having rapid reaction rates, no necessity for cofactor recycling (Taylor et al., 1998). Their relaxed substrate specificity, rapid reaction rates and no requirement for external cofactor regeneration makes transaminase enzymes attractive biocatalysts compared with chemical methods for the production of chiral amines A considerable number of à Ã¢â‚¬ °-transaminases have been identified until recently. Diamine-ketoglutaric TA is the first identified enzyme that converts the compounds bearing no carboxylic acid (Kim, 1964). It is classified as à Ã¢â‚¬ °-AT and is a member of sub-class 2 aminotransferases (Mehta et al., 1993). à Ã¢â‚¬ °-TA enzymes are known as ÃŽÂ ²-Ala:pyruvate TAs (EC 2.6.1.18) because they use pyruvate as the amine acceptor (Kaulmann et al., 2007). One of the best identified members of this group enzymes are isolated from V. fluvialis JS17. This enzyme does not show any activity towards ÃŽÂ ²-Ala but it shows broad substrate specifity towards particularly aromatic amines and (S)-enantiomers (Shin et al, 2002). It was purified and its enzymatic properties were characterised. Its molecular mass was determined to be 100 kDa and subunit mas determined to be 50 kDa. Its optiumum pH is 9.2 and optimum temperature is 37 oC. Its activity increased with pyruvate and PLP but it is inactivated with (S)-ÃŽÂ ±-methylbenzylamine. The result indicates that this is an amine: pyruvate transaminase (Shin et al., 2003). The à Ã¢â‚¬ °-AT isolated from Bacillus thuringiensis JS64 is highly enantioselective towards ÃŽÂ ±-methylbenzylamine (Shin and Kim, 1998). The à Ã¢â‚¬ °-AT Ä °isolated from Pseudomonas sp. F-126 is an isologous alpha 4 tetramer. The subunit is rich in secondary structure and consists of two domains. PLP is located in the large domain. It shows high homology with AspAT. This consequence reveals that these enzymes have common evolutionary features (Watanabe et al., 1989). In contrast to ÃŽÂ ±-transaminase catalysed reactions to produce ÃŽÂ ±-amino acids, à Ã¢â‚¬ °-transaminase reactions are not limited by a low equilibrium constant during the kinetic resolution (Shin and Kim, 1998). The enzymatic properties of three à Ã¢â‚¬ °-TAs from Klebsiella pneumonia JS2F, Bacillus thuringiensis JS64 and Vibrio fluvialis JS17 were compared to understand their mechanism and application towards production of chiral amines. All enzymes showed high enantioselectivity towards (S)-ÃŽÂ ±-MBA and broad specifity for arylic and aliphatic chiral amines. In addition to pyruvate, aldehydes showed high amino acceptor activities. All enzymes were inhibited by substrate, (S)-ÃŽÂ ±-MBA, above 200mM concentration. Only Vibrio fluvialis JS17 à Ã¢â‚¬ °-TA was inhibited by pyruvate above 10mM. The enzyme was not only inhibited by substrate but also inhibited by product. In the product inhibition case acetophenone and alanine are the main inhibitors but acetophenone is much more effective than alanine (Shin and Kim, 2001). Aminotransferases have been classified by Mehta and co-workers into four sub-groups according to their primary structure similarity (Table 1) (Mehta et al., 1993). Table 1. Classification of aminotransferases on the basis of structural similarities (Mehta et al., 1993) C:UsersadnanDesktopsub-groups of enzymes.png The members of subgroup 1 aminotransferases are Aspartate aminotransferase (AspAT), Alanin aminotransferase (AlaAT), aromatic amino acid transferase and histidine aminotransferase. Subgroup 1 aminotransferases are demonstrated to be the most versatile ones among the all subgroup hence they are able to react with alanine, dicarboxylic and aromatic amino acids. In one of the studies it was shown that the substrate specifity of AspAT and tyrosine aminotransferase overlap. This finding is based on the research which demonstrated the mitochondrial and cytosolic isoenzymes of aspartate aminotransferase from chicken heart accept L-phenylalanine, L-tyrosine and L-tryptophan as substrates (Mavrides and Christen, 1978). Group 2 aminotransferases include ornithine AT, à Ã¢â‚¬ °-AT, 4-aminobutyrate AT (GABA-AT,). They are all known as omega transaminase because of the location of the amine group. In these enzymes amino group is in a distal position from the carboxylic acid group on the amine donor substrate. (Sayer, 2009; PhD thesis). Both of the ornithine aminotransferase (Orn-AT) and 4-aminobutyrate aminotransferase (GABA-AT) are pyridoxal-phosphate (PLP)-dependent enzymes that have been identified in human, plants and animals until recently. Both enzymes catalyse a wide range of reactions on amino acids (Storici et al., 1999; Markova et al., 2005). Each enzyme catalyses the transamination reactions by a ping-pong bi-bi mechanism. The mechanism comprises two-half reactions. The half-reaction converting ketoglutarate to glutamate is the same for all transaminases. Therefore, the change in substrate specifity is resulted from the second half reaction in which an amino group is transferred distant from the ÃŽÂ ±-carbon. As a result of this, these enzymes have been identified as omega transaminases (Markova et al., 2005). GABA aminotransaminase is a PLP dependent and Fe-S cluster containing enzyme which involves in regulation of the concentration of major inhibitory neuro-transmitter GABA. This enzyme degrades GABA to succinic semialdehyde (Storici et al., 1999). Subgroup 3 aminotransaminases are BcaaAT and D-alanine aminotransferase (DaAT). The amino acid substrates of the two members of subgroup 3 enzymes have different chirality but they share the same oxo-acid as substrate. Subgroup 4 aminotransferases include SerAT and pSerAT. The two members of this group act on structurally and biosynthetically related substrates (Mehta et al., 1993). Grishin and co-workers further classified all PLP-dependent enzymes and aminotransferases categorizing them according to their PLP folds (Grishin et al., 1995). Similar outcomes were obtained with Mehtas classification. One of the classifications has been performed by John Ward in which he classified à Ã¢â‚¬ °-ATs in four groups according to their substrate specifity. 1. ÃŽÂ ²-alanine:ÃŽÂ ±-ketoglutarate aminotransferase, highly specific substrate activity. 2. MBA (ÃŽÂ ²-alanine):pyruvate aminotransferase, broad substrate specificity. 3. MBA:pyruvate aminotransferase, broad substrate specificity but inactive on ÃŽÂ ²-alanine. 4. ÃŽÂ ²-alanine (MBA):pyruvate aminotransferase, broad substrate specificity.(Sayer, 2009; PhD thesis). The à Ã¢â‚¬ °-AT from Chromobacterium violaceum belongs to the sub-group 3. The research which was conducted by Kaulmann and co-workers demonstrated that his enzyme does not show any activity towards ÃŽÂ ²-alanine where as it has comparatively broad substrate specifity against aromatic, aliphatic amines and amino-alcohols. It has a molecular weight of 51 kDa and shows 38% sequence identity to the à Ã¢â‚¬ °-AT from V. fluvialis JS17 (Kaulmann et al., 2007). PLP-DEPENDENT ENZYMES PLP is an important cofactor for amino acid metabolism. PLP-dependent enzymes catalyse a wide range of reactions such as transamination, decarboxylation, racemisation, aldol condensation, ÃŽÂ ±,ÃŽÂ ²-elimination and ÃŽÂ ²,ÃŽÂ ³-elimination of amino acids, and amine oxidation (Soda et al., 2001). PLP forms a covalent bound with the substrate molecule and performs as an electrophilic catalyst (Percudani and Peracchi, 2003). The mechanistic studies revealed two key chemical characteristics of the cofactor; an imine is formed between aldehyde group of PLP and amino group of substrates. The other basic characteristic of the cofactor is being able to perform as electron sink and withdrawing electron from substrate compounds (John, 1995). In 1974, it was hypothesised that the complete family of PLP-dependent enzymes had evolved from a common ancestor. They proposed this hypothesis the result of the investigating of the mechanism of the seven PLP-dependent enzymes. It was observed that protonation of the C4 carbon of the coenzyme proceeds stereospecifically with the same stereo-face in all different enzymes. This result is is explained as proof for the evolution of complete family of PLP-dependent enzymes from a common ancestorial protein (Dunathan and Voet, 1974). REACTION MECHANISM OF TRANSAMINASES The aminated form of PLP, pyridoxamine 5à ¢Ã¢â€š ¬Ã‚ ²-phosphate (PMP), appears only in the transamination reactions (Fig. 1). Transamination reactions basically comprise of two half reactions. In the first step, the aldimine is deprotonated to be converted into a quinoid intermediate, which in turn accepts a proton at a different position to form a ketimine. The resulting ketimine is hydrolysed leaving PMP behind which performs as an amine donor in the second half-reaction. PLP is subsequently recycled. PMP interacts with the apoenzyme via only non-covalent interactions, whereas, PLP is covalently bound to the active site lysine residue of the enzyme molecule. Apotransaminases have been identified to bind PMP about 100-fold less tightly than PLP. PMP can be displaced from the enzyme with high concentrations of sulphate or phosphate ions (Schell et al., 2009). C:UsersadnanDesktop.gif Scheme 1. The first half reaction mechanism of aminotransferases (Adapted from Schell et al., 2009). STRUCTURE OF the Chromobacterial OMEGA TRANSAMINASE The C. violaceum à Ã¢â‚¬ °-AT is the first enzyme among à Ã¢â‚¬ °-aminene pyruvate AT whose structure was investigated in detail using X-ray. The C. violaceum à Ã¢â‚¬ °-AT protomer is folded into two domains similar to other class II aminotransferases that were classified by Mehta and co-workers based on their primary structure similarity. The enzyme comprises of a large domain which includes residues 62-343 and a relatively small domain containing the N and C-terminal parts of the polypeptide chain residues between 6-61 and 344-456. The large domain has a typical ÃŽÂ ±/ÃŽÂ ²/ÃŽÂ ± sandwich fold constituted a central seven stranded ÃŽÂ ²-sheet and helix-loop-helix segment. The small domain is constituted largely of the C-terminus which is comprises of ÃŽÂ ²-sheet that is packed against helices. The overall protein fold is represented in figure 1 (Sayer, 2009; PhD thesis). Figure 1. The tertiary structure of the C. violaceum à Ã¢â‚¬ °-AT promoter. The ÃŽÂ ²-strands are tagged as S, ÃŽÂ ±-helices are marked as H (Sayer, 2009). As it was mentioned previously the aminotransferases require PLP as a cofactor. The enzyme was crystallised with PLP to investigate the binding interactions of PLP with active site residues. The cofactor is linked to the active site lysine residue through covalent bond forming lysine-pyridoxal-5-phosphate in all four sub-units. The PLP binding site is demonstrated to be situated between small and large domains at the interface of the two sub-units. The oxygen atoms of phosphate moiety interact with the main-chain amides of Gly120, Ser121 and the side chain of Ser121. The carboxyl group of Asp259 is located within hydrogen bond distance to the pyridine nitrogen of PLP. Aspartic acid is interacted with Val261 and Histidine154. The interactions are demonstrated in figure 2 (ibid). plp_contacts Figure 2. Stereo diagram of the C. violaceum haloenzyme active site. Stick model represents the lysine-PLP Schiff base. Yellow dot lines represent hydrogen bonds and the interacting residues are shown as lines. Neighbouring subunit residues are demonstrated by *. The structure of C. violaceum à Ã¢â‚¬ °-AT gabaculine complex is also solved. Gabaculine is a naturally occurring inhibitor and first isolated from Streptomyces toyacaensis as an inhibitor of GABA-AT (Kobayashi et al., 1977). Gabaculine interacts to the aminotransferase forming Schiff base with PLP as the m-carboxyphenylpyridoxamine phosphate (mCPP) ligand in the subunit A. The inhibitor molecule is interacted to the enzyme on the re face of the cofactor at the bottom of the active site. A salt bridge is formed between carboxyl group of gabaculine and side chain of Arg416. Despite the fact that à Ã¢â‚¬ °-AT act on the substrates that do not contain carboxyl groups, amine derivatives that bear a carboxyl group will be orientated by Arg416. The gabaculine is surrounded by Trp60, Ala231, Ile262, Leu59 and His318 from the neighbouring subunit to the bound cofactor. The hydrophobic pocked is formed through this interaction.. The interaction between active site residues and m CCP is in dicated in figure 3. gaba_lines_stereo Figure 3. Stereoview of the C. violaceum à Ã¢â‚¬ °-AT bound with gabaculine in the presence mCPP ligand (represented with stick) and amino acid residues within 4.5 Ã…. Residues from the neighbouring subunit to the bound cofactor are demonstrates as *. The structure of C. violaceum à Ã¢â‚¬ °-AT with pyruvate-PLP complex was solved to investigate active site pyruvate binding pocket (Figure 4). The carboxyl group of pyruvate forms a salt bridge with Arg416 and a hydrogen bond to the indole nitrogen of Trp60. The overall pocket is hydrophobic made up by the residues Tyr168, Phe22, Phe88, Leu59, Ala231 and Ile262. pyr_lines Figure 4. Stereoview of the binding interaction of the C. violaceum à Ã¢â‚¬ °-AT with pyruvate-PLP complex is demonstrated as stick model. Hydrogen bonds are represented in yellow colour, * shows residues from the adjacent subunit to the bound cofactor. BIOTECHNOLOGICAL APPLICATION OF OMEGA TRANSAMINASES A number of important parameter such as enantioselectivity, reaction equilibrium stability of enzyme, effect of inhibitors, and product separation must be taken into account in order to perform successful kinetic resolution and asymmetric synthesis for the synthesis of enantiomerically pure amines (Kim et al., 2003). Enantiomerically pure amines can be synthesized using two fundamental methods that employ à Ã¢â‚¬ °-ATs. One of them is kinetic resolution that performs on racemic amines; the other one is asymmetric synthesis starting with prochiral ketones that together correspond to the transamination reaction run forward and in reverse subsequently. Amines with opposite conformation are attainable if the same omega transaminase in kinetic resolution or asymmetric synthesis. For instance, if (S)-enantiomer is observed during asymmetric synthesis, the same enzyme will produce the (R)-enantiomer in the kinetic resolution (Koszelewski et al., 2010). Despite the fact that, asymmetric synthesis permits a 100% yield of demanded optically pure compound, it has been preferred less in recent time due to the difficulties related to reaction equilibrium and stereoselectivity. The stereoselectivity of the enzyme molecule to produce amines must be high with ee value of >99%, but it is almost impossible for the available à Ã¢â‚¬ °-transaminase (ibid). Kinetic Resolution In some reaction an enzymatic reaction is performed between a chiral molecule and a racemic acid mixture. In this case kinetic resolution occurs resulting to a kinetic preference, for one of the enantiomer over the other enantiomer (Novasep, 2010). The significance of kinetic resolution has remarkably increased as the importance of optically pure amines has increased. One group of enzymes that have resulted in the production of optically active compounds are transaminases. One of the simplest techniques for investigating kinetic resolution of chiral primary amines involves the employment of a stoichiometric equivalent of the amino acceptor. In this approach, the thermodynamic equilibrium is on the product side and comprises the enantiomerically rich amine, ketone and amino acids. The main positive side of this method is that it needs only à Ã¢â‚¬ °-AT (Koszelewski et al., 2010). As it is mentioned previously this method has been used commonly but it has two main drawbacks that ketone product and starting pyruvate molecule cause an inhibitory effect on the à Ã¢â‚¬ °-TAs enzymes (Yun et al., 2005). Different strategies have been developed to overcome these problems. The à Ã¢â‚¬ °-TA from Vibrio  ¬Ã¢â‚¬Å¡uvialis JS17 has been identified to show high enantioselectivity for the (S) enantiomers of various chiral amines, such as ÃŽÂ ±-MBA and sec-butylamine, with remarkable stability and a high reaction rate (Shin and Kim, 1998). Nevertheless, production of ketone may result in the inhibition of the enzyme preventing it to show its activity (Shin and Kim, 1997). A restricted solution to this issue was using an extractive biphasic reaction system that reduced the inhibitory acetophenone concentration in the aqueous phase (ibid). However, in this system the organic phase interacts with aqueous phase and acts as extractant of acetophenone. To keep the concentration of acetophenone in the aqueous phase at low levels, the aqua phase was unavoidable because the exchange the organic extractant need to control the aqueous pH with acid to neutralise basic ÃŽÂ ±-MBA diffused from the organic phase also made the process complex. An enzyme-membrane rea ctor (EMR) coupled with hollow-fiber membrane contractor was employed for the production of chiral amines and to get over the problems in a two-liquid phase reaction system. In the EMR system, to preserve the extraction capacity, a simple exchange of solvent in the organic reservoir and pH control are necessary to transfer only acetophenone through the contractor. Other advantage of this system over two-liquid phase reaction system is confining the enzyme in the reactor with ultrafiltration membrane eases reuse of enzyme. The main problem with this system could possibly be the economics of the availability of purified enzyme because a sufficiently high-circulation rate of sunstrate solution is desirable to residence time and minimize the product inhibition (Shin et al., 2001). An effective synthesis of enantiopure (S)-amino acids and chiral (R)-amines was carried out using ÃŽÂ ±/à Ã¢â‚¬ °-AT coupling reaction and à Ã¢â‚¬ °-AT was found to be inhibited by ketone product. To remove inhibitory reaction product a two-liquid phase reaction system in which dioctylpthalate was selected as solvent to achieve the best system. One of the most important advantages of ÃŽÂ ±/à Ã¢â‚¬ °-AT coupling reaction over aqueous phase is that; it can be carried out at high substrate concentrations to fulfil industrial large scale production of chiral amine and amino acid compounds (Cho et al., 2003). A concept has been developed to improve rate and enantioselectivity in à Ã¢â‚¬ °-AT-catalysed kinetic resolution using a protection group. For this purpose the kinetic resolution of 3-aminopyrrolidine and 3-aminopiperidine with à Ã¢â‚¬ °-AT was expedited using a protective group. 1-N-Cbz-protected group. Upon application of protective group the reaction rate was 50-fold higher. Enantioselectivity was also considerably increased upon carbamate protection in comparision with the unprotected compound (86 vs.99 ee%). However, benzyl protection of former substrate did not affect enantioselectivity because of the difference in the flexibility of the benzyl- or carbamate-protected 3-aminopyrrolidine. Despite of 50% yield limitation in kinetic resolution, this strategy is an efficient way to synthesise enantiopure 3-aminopyrrolidines (Hà ¶hne et al., 2008). The other approach to overcome product inhibition by aliphatic ketones is using an enrichment culture in combination with random mutagenesis for production and purification of mutant à Ã¢â‚¬ °-TA. This technique is mainly based on using 2-aminoheptane as amine donor and nitrogen source in minimal medium, and 2-butanone as an inhibitory ketone. Consequently, the higher growth rates of mutants resistant to inhibition allow them to be enriched in culture reducing the number of colonies that needs to be screened. A mutant enzyme, à Ã¢â‚¬ °-TAmla, which shows significantly reduced product inhibition by ketone, was determined. Using this mutant enzyme 2-aminoheptanone was resolved to (R)-2-aminoheptane with ee value>99, 53% conversion and enantioselectivity of >100 (Yun et al., 2005). b) Asymmetric synthesis Performing an asymmetric synthesis reaction is not as easy as kinetic resolution reactions because of unconvenient equilibrium and product inhibition. The main advantage of asymmetric synthesis over kinetic resolution is resulting 100% yield in the production of desired optically pure amine. However, side products may affect the enzyme catalysed reaction; therefore, these problems must be overcome to apply successful asymmetric synthesis (Koszelewski et al., 2010). One of the easiest methods for amination involves applying an excess of amine donor due to the necessity of only a single transaminase. Nevertheless, the issue here is the reaction equilibrium and potential inhibition by co-product and excess of starting an amine. In one of the studies, alanine was applied in 16-fold excess for the amination of 4-methoxyphenylacetone with 94% conversion (Nakamichi et al., 1990; Koszelewski et al., 2010). To overcome pyruvate inhibition problem two enzyme system has been used. One of the commonly used method involves Lactate Dehydrogenase (LDH)-reduction of pyruvate. One-pot, two-step dereacemisation cascade reaction was employed to lead to the production of optically pure pharmaceutical intermediates through kinetic resolution and following stereoselective amination. The main advantage of this cascade reaction is circumventing the restriction of kinetic resolution (50% conversion) leading high yield of optically pure amines. In the second step, side product pyruvate was removed using lactate dehydrogenase to shift the equilibrium to the product side. The disadvantage of this system is requirement for coenzyme recycling (Koszelewski et al., 2009). The use of whole cells is hindered by the reason that undesired side reaction such as the reduction of alcohol to ketone products. In one of the studies it was shown that the equilibrium can be shifted using pyruvate decarboxylase (PDC). Dec arboxylation of pyruvate to produce acetaldehyde and CO2 with PDC is more advantageous than LDH-catalysed reduction of pyruvate owing to no requirement for cofactor recycling (Hà ¶hne et al., 2008). One of the important examples of asymmetric synthesis is the amination reaction of acetophenone with alanine for the objective of producing (S)-ÃŽÂ ±-MBA. The equilibrium constant of the reaction is 8.8110-4 and (S)-ÃŽÂ ±-MBA and pyruvate are more reactive substrates than acetophenone and alanine (Shin and Kim, 1999). The stereoselectivity of C. violaceum TA-mediated amination of an ÃŽÂ ±,ÃŽÂ ±-dihydroxyketone, 1.3-dihydroxy-1-phenylpropane-2-one, was investigated. It was shown that the enzyme is not enantioselective towards the racemic 1.3-dihydroxy-1-phenylpropane-2-one, whereas; it is highly stereoselective for the (2S)-2-amino-1-phenyl-1,3-propanediols in 99% ee (Smithies et al., 2009). CONCLUSION AND AIMS OF PROJECT After drawing various analyses, the general overview, structure, mechanism and biotechnological applications of à Ã¢â‚¬ °-transaminases were reviewed in order to shed some lights on the characteristics of the à Ã¢â‚¬ °-transaminases. Transaminases have been used broadly owing to its capacity to produce amino acids and chiral compounds which are important for pharmaceutical industries. Transaminases have been divided to sub-groups according to their substrate specifities and structure. Transaminases, under sub-group 2 are known as à Ã¢â‚¬ °-AT because the distal amino group of the substrate undergoes the reaction and include OrnTA, GABA-TA and à Ã¢â‚¬ °-amino acid:pyruvate AT. Among them the most important one is à Ã¢â‚¬ °-amino acid:pyruvate AT due to the reason that only this enzyme shows catalytic activity towards primary and aliphatic amines bearing no carboxyl group. The other advantages of à Ã¢â‚¬ °-TAs are having broad substrate specifity, high enantioselectivity an d no requirement for the cofactor recycling. Two main biotechnological applications of à Ã¢â‚¬ °-ATs are asymmetric synthesis and kinetic resolution. They have both advantage and disadvantage over each other. The disadvantages of kinetic resolution are being subjected to pyruvate and ketone inhibition and having 50% yield. The asymmetric synthesis results in 100% yield for the manufacturing of desired optically pure amine. However, product may cause to the inhibition of the enzyme. To overcome these problems some approaches have been developed including biphasic reaction system, using enzyme-membrane reactor (EMR) coupled with hollow-fiber membrane contractor, using protective groups. The aim of this project is to purify and crystallise the à Ã¢â‚¬ °-AT enzymes and characterise their subsrate specifity.

BonJours Basic Antifoundationalist Argument :: Philosophy Philosophical Papers

BonJour's 'Basic Antifoundationalist Argument' ABSTRACT: BonJour argues that there can be no basic empirical beliefs. But premises three and four jointly entail ‘BonJour’s Rule’ — one’s belief that p is justified only if one justifiably believes the premises of an argument that makes p highly likely — which, given human psychology, entails global skepticism. His responses to the charge of skepticism, restricting premise three to basic beliefs and noting that the Rule does not require ‘explicit’ belief, fail. Moreover, the Rule does not express an epistemic duty. Finally, his argument against this fails since it is false that if an experiential state has representational content, then it is in need of justification. I venture the diagnosis that BonJour mistook the representational content of a cognitive state for the assertive functional role of a belief. Foundationalism may well be false, but not for BonJour’s reasons. Laurence BonJour observes that critics of foundationalism tend to argue against it by objecting to "relatively idiosyncratic" versions of it, a strategy which has "proven in the main to be superficial and ultimately ineffective" since answers immune to the objections emerge quickly. (1) BonJour aims to rectify this deficiency. Specifically, he argues that the very soul of foundationalism, "the concept of a basic empirical belief," is incoherent. (2) This is a bold strategy from which we can learn even if, in the end, as I shall argue, it fails. But, first, what is foundationalism? A person's belief is ‘nonbasic’ just in case it is justified in virtue of its relation to other justified beliefs; it is ‘basic’ just in case it is justified but not in virtue of its relation to other justified beliefs. Foundationalism is the view that if one has a nonbasic belief, then — in the final analysis — it is justified in virtue of its relation to a basic belief. Basic beliefs comprise the foundation of a person's network of justified beliefs. Now to BonJour's argument. 1. The Argument Stated BonJour summarizes it like this: 1. Suppose, for reductio, that there are basic empirical beliefs. 2. A belief is justified only if there is a reason why it is likely to be true. 3. A belief is justified for a person only if he is in cognitive possession of such a reason. 4. A person is in cognitive possession of such a reason only if he believes with justification the premises from which it follows that the belief is likely to be true.

Prejudice Against Native Americans Essay -- essays research papers

Prejudice Against Native Americans These people began migrating thirty thousand years before Christopher Colombus "discovered" the Americas. Native Americans migrated from Asia, crossing a land bridge where the Bering Strait off the coast of Alaska is today. Over the centuries these people spread throughout the continents of North and South America. Since the arrival of the Europeans in 1492 the American Indian has been dehumanized, decivilized and redefined into terms that represent a dominate European view. The Spanish explorers under Colombus were the first to use the terms "Indian" to mean a Native American. These explorers were under the false impression that the had reached the West Indies. This term is still used today. From the first interaction with the native peoples the Europeans inatiated dominance and superiority. There are three distinctive reasons that the Europeans were able to dominate and later oppress the Native American culture such as; the Native American relgious beleifs and practices, the lack of interaction between Native Americans ans Europeans and the lack of orginization of the Indian tribes. All of these aspects had a strong influence the Europeans to become dominate figures on the Native American land. These factors can still be attributed for the way that Native Americans are viewed in society today. After the Revolutionary War the new United States government sought to gain land through treaties. Th...

Animals and Nature in the Work of Margaret Wise Brown :: Biography Biographies Essays

Animals and Nature in the Work of Margaret Wise Brown Read almost any book by Margaret Wise Brown, and you will start to see some overlapping trends. Readers know when they are reading a work by this famous author without seeing the cover or title page because her works have so many similarities. The use of multiple animals and nature frequently appear in her books and serve as common ideas in literature by Margaret Wise Brown. Many of Margaret Wise Brown’s most famous books have animals as the main character. For example, Runaway Bunny and Goodnight Moon, two of her most popular books, feature rabbits as the main characters. Further, in Goodnight Moon, the animal’s behavior is parallel to that of humans. For example, the motherly figure on the rocking chair is reading to the young rabbit as many parents do to their children. Additionally, Brown adds a humorous element as the young rabbit seems to have pets. Margaret Wise Brown was truly fascinated by animals, and she understood children’s attraction to animals. Tellingly, when Brown reflects on her childhood she mentions her â€Å"thirty-six rabbits, two squirrels†¦a collie dog, and two Peruvian hens, a Belgian hare, seven fish, and a wild robin who came back every spring† (Days Before Now). From this information about Brown, one understands where her love of animals originated--her childhood. Additionally, animals were kind to her and did not restrict or belittle Brown the way some individuals did regularly. Brown was allowed to have constant interaction with animals, which proved to be influential in her writing career. Overall, Margaret Wise Brown used numerous animals, especially rabbits because of her love for creatures and the understanding she possessed of children’s love of and interest with animals. Another area of focus when creating her books is nature. As a child, nature became Brown’s life as she notes, I grew up along the beaches and in the woods of Long Island Sound. This was the country. And from then on I was terribly busy hitching up all the dogs I could find to pull me around on my sled in the snow, and picking cherries high up in cherry trees, chasing butterflies, and burning leaves, and picking up shells on the beach, and watching the new flowers come up in the woods as the seasons passed (Days Before Now) This passage gives readers an enhanced understanding of this talented author, as they see her passion for the wilderness during childhood.

Thesus vs Achilles

Dumbledore said, â€Å"It is the choices†¦ that show what we truly are, far more than our abilities. † This theory applies to everyone. Through the choices and actions of Theseus and Achilles, we learn a lot about their characters. We find Achilles to be motivated by rage and pride, with ambitions of glory and vengeance. Theseus, on the other hand, seems to be motivated by others’ needs, and aspires to do right by those who he should help. Theseus also likes to challenge expectations and overcome the â€Å"impossible†. Theseus begins his saga with a journey to Athens. He chooses the difficult and dangerous land route, as opposed to the sea route, because he wants the challenge. On his way, he completes six tasks that make him a great hero. He kills robbers and murderers, helping innocent potential victims, delivering justice, and making a name for himself. Once he arrives in Athens, he kills some of the people who plan to attack him. Even though they are not innocent and pose a threat to him, he doesn’t kill about half of them, including his uncle Pallas. This shows his solid reasoning. When Theseus captured the bull of Marathon, he met a normal, old lady, and paid her significant respects in her death. Theseus was seen as a â€Å"champion of the oppressed. † For example, he gave Oedipus a place to live after he chose exile. Achilles’ ambitions and motivations had little in common with Theseus’. He was driven by anger, the need for vengeance, bloodlust, and excessive pride. He strived for eternal glory. Achilles played a huge role in the Trojan War, a situation that aligned perfectly with his personality. He showed no humility in slaying others. He mutilated Hector’s body. He refused to accept Agammemnon’s great gifts out of his hubris. In the end, his need for vengeance overcame even his dispute with Agammemnon, and his sole purpose was to destroy Hector, afterward mutilating his body. He was miserable, but finally learned compassion. Achilles and Theseus were very different heroes. Their choices and actions clearly outline their motivations and ambitions for us. Theseus is motivated by the excitement of challenges, and the opportunity to help others. His ambitions are greatness, but, more importantly, justice and aiding those in need. Achilles’ motivations are negative and his ambitions shallow. However, at the end of his life, we see a positive change in his personality.

Benefit of changing to Activity Based costing system

Activity Based Costing has been of great help to the companies and other forms of businesses. With the help of ABC, the organizations have been able to implement strategies to overcome various problems facing the welfare of their businesses and other activities in the organizations. The organizations have been able to assign cost of each activity to the products and services that are offered by a particular organization. This is done for the sole purpose of elimination of high prices to some products and due to lack or low profits in other products or services.The ABC rule is used as a tool by businesses to assign the cost of the business activities to the goods and services (organization products) offered by the business to its customers. It is used as a tool that understands the customers and the product cost and the overall profitability. It is also used to in supporting the decision making process as would be used in product pricing, marketing, identification and a way to measure the initiative of improvements. IntroductionThe Activity-Based Costing (ABC) is a method used by accountants that can allow the business to collect data in regard to their operations costs. Their operations cost is then assigned to specific activities that include planning, production and other organization activities and then they are associated with the variety of products and services. This enables the company to know what products and services have increasing profits and which have increased losses, this help in learning the flow of each product and service within the company.The use of ABC method help the concerned personnel in the company to prepare an effective data that will aid in preparing budget that will bring a greater achievements in the company and enable the company to have an effective plan for the expense that will keep the company running smoothly for a given period of time. ABC method is mostly effective and is much recommended for a longer period of time over t he short term as opposed by the theory of constraints. Research objectives The objectives of this research are: †¢ To determine the effectiveness of ABC†¢ To investigate how companies implement the ABC in their activities †¢ To compare the excellence of companies that use ABC and those that don’t use ABC †¢ To determine the advantages and disadvantages of ABC Literature review The activity based costing got first implementation in the 1980s. It was a formed as an alternative to replace the traditional way of cost management systems that was not effective in producing favorable results when allocating the costs. The implementation of the system was advocated by Professor Robert Kaplan of Harvard business school.It was initially used for private business though it has currently gained usage by even the public forums like those used to measure the efficiency of the governance. The ABC programs require proper planning and great commitment from the highest mana gement. It is recommendable to start up with trials in the organization. These trials may be done with the departments that are not performing well as it would be expected of them. This would be a great chance to show that the ABC is an effective system that would save money for the company with great achievements.On the other hand, if the company des not save money on this, either the system process was not well implemented or may not be effective for the company. (Gary, 1998) According to the study, the business should first set up a team that will master the progress of the product and which activity would be appropriate for the product and service. The team should include all persons from the different departments of the company. After the formation of the team and putting in place all the required materials, the activities that cost money are determined. (Cokins, 1999 and David 1991)At this point, great attention is paid for details on each activity are required, most of the co sts might be hidden and may bring in controversy there after for the may not be obvious initially. Even though there might be some replacement to human labor, it’s still important to determine the fixed cost of a product or service. (Cokins, 2000) Even though this method is important to the business, it has some risks. The companies that use the ABC system, the risk using too much time and money while trying to implement the system and collecting data, going through it and in implementations.Lack of detailed data can lead to frustrations of the managers and can on the other hand contribute to the down fall of the system. Mistake done on the data collection can lead to downfall again if not noticed before implementation. (Joyce, 1999) Through the implementation of the activity based costing, the employees are able to understand the different costs that are involved in different activities. This enables them to analyze and identify the activities that add value to the product a nd those that do not, it helps them improve on value and benefits are realized.In the DHL Company, the employees are involved in collecting data and views from the clients; this enables them to involve themselves in quality improvement of their services and eliminate the non profitable activities that enable the overall company’s efficiency. (Henricks, 1999) The ABC system has helped the DHL Company to answer the market need in providing quality service at a better and competitive price. This system has helped the company’s to management in the decision making process. (Hicks, 1998) This has helped the company to reduce the price of their services without a decline in value of their services.(Hicks, 1999) The benefits of the ABC system are now in use in most institutions that include the government departments and the public financial institutions. The Barclays bank New York changed its accounting method to provide accurate and cost effective data. Through the change t o ABC, the Barclays bank has particularly found it helpful compared to the former funding based accounting systems that was designed for ensuring legal compliance. (Paulo, 1998) In the use of ABC, the managers are able to direct funds where they are more beneficial.They are able to learn the weaker points in the organization making it easy for them to implement the most beneficial activity in the business. (Robert, 2004) The managers in Barclays bank found that ordinary accounts were not doing much well compared to loans, they campaigned for people to take more loans at simpler rates and prolonged repayments period, and this increased to Banks profits than ever. The bank is launching new loan strategies that are expected to double the profits. (Robert and Bruns, 1987)