cDNA cloning, overexpression in Escherichia coli, purification and characterization of sheep liver cytosolic serine hydroxymethyltransferase

A sheep liver cDNA clone for the cytosolic serine hydroxymethyltransferase (SHMT) was isolated and its nucleotide sequence determined. The full-length cDNA of SHMT was placed under the control of T7 promoter in pET-3C plasmid and expressed in Escherichia coli. The overexpressed enzyme, present predominantly in the soluble fraction, was catalytically active. The recombinant SHMT was purified to homogeneity with a yield of 10 mg/1 bacterial culture. The recombinant enzyme was capable of carrying out tetrahydrofolate-dependent and tetrahydrofolate-independent reactions as effectively as the native enzyme. The Km values for serine (1 mM) and tetrahydrofolate (0.82 mM) were similar to those of the native enzyme. The recombinant enzyme had a characteristic visible spectrum indicative of the presence of pyridoxal 5′-phosphate as an internal aldimine. The apoenzyme obtained upon removal of the cofactor was inactive and could be reconstituted by the addition of pyridoxal 5′-phosphate demonstrating that the recombinant SHMT was functionally very similar to the native SHMT. This overexpression of eukaryotic tetrameric SHMT in E. coli and the purification and characterization of the recombinant enzyme should thus allow studies on the role of specific amino acids and domains in the activity of the enzyme

Preliminary Survey on the Crucial Root Causes of Material Waste Generation in Malaysian Construction Industry

Construction waste generation has been perceived as a crucial issue that has critical consequences on the project effectiveness and ecological effect of the construction industry. The key objective of the current research is to identify the root causes of material construction waste generation in Malaysia. A well-structured questionnaire was designed based on the 52 identified root causes of material waste generation. The developed questionnaire was distributed to 35 highly experienced in Malaysian construction industry experts. The collected data was analyzed by the use of mean score analysis. The top identified root causes are poor supervision during the construction stage, lack of efficient site management, last minute changes, cutting uneconomical sizes of materials, and constant design changes during the construction period. The initial findings will aid construction practitioners to focus on these root causes in order to reduce the material construction waste generation at construction sites

Anomeric Effect in Phosphate Moieties of Carbohydrates: An ab initio Study

Ab initio SCF MO calculations at the STO-3G level have been carried out on $CH_2OHOPO_3^-H$ and $CH_2OHOPO_3^{2-}$, which have been considered as model systems for the C1-phosphate moiety of sugars. The results predict higher anomeric energy for the phosphate moiety at C1 atom of pyranosides. Also a trans arrangement of the exocyclic 0-P bond is preferred rather than a gauche arrangement, thus exhibiting a reverse exo-anomeric effect. A complete potential energy map has been constructed for $CH_3OP_3^-H$, a model system for C6-phosphate moiety. It is seen that the bond angle optimization brings down the relative energies of various conformations. The effect of the phosphate group on the preferred conformation of phosphate containing polysaccharides is also discussed

Anomeric effect in carbohydrates-an ab initio study on extended model systems

The anomeric effect is studied at the ab initio Hartree-Fock level using extended model systems. This has enabled us to study the effect of substitution on the anomeric effect and the $\Delta ^2$-effect in greater detail. An attempt is made to compare the results with available experimental data

Ab initio SCF-MO study of the molecular structures of aminomethanol, aminesulfonic acid and N-methyl-sulfamate

Ab initio MO studies at the STO-3G and the 4-31G level are reported for aminomethanol (H<SUB>2</SUB>NCH<SUB>2</SUB>OH), aminesulfonic acid (H<SUB>2</SUB>NSO<SUB>3</SUB>H) and N-methylsulfamate (CH<SUB>3</SUB>NHSO<SUB>3</SUB><SUP>-</SUP>). The potential energy surface of aminomethanol is compared with that of methanediol and the origin of "reverse anomeric effect" is discussed. The potential energy surface of aminesulfonic acid is compared with that of aminomethanol and sulfuric acid. The favored conformations are analysed in terms of the interaction of polar bonds and the dipoles on the atoms. Various contributions to the potential energy are analysed by decomposing the potential into three term Fourier components. A general survey is made of the effect of bond length and bond angle optimisation on the conformational energies of various anomeric systems

Ab initio SCF-MO study on difluoromethanediol and difluorodimethoxymethane

Difluoromethanediol and difluorodimethoxymethane systems have been investigated by ab initio molecular orbital methods using STO-3G and 4-31G basis sets. The potential energy surfaces for both systems have been constructed and compared with those of methanediol and dimethoxymethane, respectively. Difluoromethanediol favours (180&#176;, 0&#176;) as the minimum energy conformation whereas difluorodimethoxymethane favours (180&#176;, 60&#176;). However, after bond angle optimization difluorodimethoxymethane favours (180&#176;, 180&#176;) conformation. A comparison of the minimum energy conformations of phosphates, sulfates and methanediol with that of difluoromethanediol has been made. Bond length optimization reveals very short C---F and C---O bonds in difluoromethanediol. Decomposition of the potential function V(&#248;1) using a three-term truncated Fourier expression shows that the V1 term becomes less important in difluoromethanediol, when compared with the methanediol system

Theoretical investigation of the structures of H<SUB>2</SUB>SO<SUB>4</SUB>, (CH<SUB>3</SUB>O)<SUB>2</SUB>SO<SUB>2</SUB> and CH<SUB>3</SUB>OSO<SUB>3</SUB>

The energies of the various conformers formed by rotating around the S---O bond in H2SO4, (CH3O)2SO2 and CH3OSO3- have been determined by ab initio molecular orbital methods using minimal (STO-3G) and extended (4-31G) basis sets. The (90&#176;, 90&#176;) conformation in H2SO4, has been found to be the most stable, in agreement with the microwave spectral data for gaseous sulfuric acid. Calculations on dimethyl sulfate indicate that geometry optimization can dramatically alter the relative energies of the conformational isomers. Thus, after RO---S---OR bond angle optimization, the energy difference between the (75&#176;, 75&#176;), (75&#176;, 180&#176;) and (180&#176;, 180&#176;) conformations is only about 0.21 kcal mol-1 suggesting that all three rotational isomers are likely to occur. This explains the detection of different conformers of dimethyl sulfate by different experimental techniques. The barrier to rotation around the S---O bond in the CH3OSO3- system is calculated to be very small, &#8764;1.2 kcal mol-1. A comparison of these species with related anomeric systems is presented and the nature of the interactions of polar bonds in determining the preferred conformation is discussed

A review of SMEs recruitment and selection dilemma: Finding a fit

This systematic review of theoretical approaches is to determine whether the recruitment and selection process in SMEs shows evidence of a strategic &#039;fit&#039; and second, examining the extent of a structured, innovative and flexible recruitment and selection process. HR managers may be aware of Strategic Human Resource Management but their understanding of recruitment and selection process as a HR architecture linking the firm and HR strategy may be vague. Findings in literature shows that priority research tends to be non-HRM related functions. HRM practices are seen as less important. The review also shows evidence that recruitment and selection is an under researched area and no specific research papers provided insights on the concept of &#039;fit&#039; in recruitment and selection. Although general literature presents various models of recruitment but the compatibility between the individual, job and environment - &#039;fit&#039;, as a strategic process in SMEs seems disconnected and unexplored. This suggests that the HRM functions in the Malaysian SMEs are treated and perceived as secondary, routine and an insignificant function