Thesis (Ph. D.)--Massachusetts Institute of Technology and Woods Hole Oceanographic Institution, 1985. / Bibliography: p. 434-451.

A 5' cDNA clone coding for human C4b-binding protein (C4bp) was isolated, characterised and sequenced to complete the cDNA sequence coding for amino-acid residues 1-32 thus confirming the protein sequence data of Chung et al. (1985c). The sequence extended to allow derivation of the putative leader sequence which was 32 residues in length and showed a high degree of hydrophobicity typical of other documented leader sequences. This clone together with others isolated by Chung et al. (1985a) were used to prepare cDNA probes which were subsequently used to hybridise genomic clones. The data from the blots suggested that the C4bp gene was up to 30 kb in size and thus consisted of large proportions of intron sequence (mRNA ≃ 2.5 kb). A commonly occurring restriction fragment length polymorphism was detected using the BglII restriction enzyme. Analysis of the genomic clones has shown that, with one possible exception, each internal protein repeat is encoded by a discrete exon. Use of the cDNA probes to hybridise RNA transfer blots, on which the RNA originated from various tissues and cell-lines, suggested that the liver is the major source of C4bp mRNA. Cross hybridisation was detected between the human C4bp cDNA probes and genomic DNA isolated from various species on transfer blots suggesting that genomic sequence homologous to that coding for C4bp has been conserved during evolution. Results from pulse field gel electrophoresis blot analysis suggests that although C4bp and the alternative pathway regulatory protein, Factor H, are genetically linked the physical distance separating them may be as great as 3 Mbp.

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No description available.

Clostridium perfringens was found to produce marked changes in its environment during growth. Decrease in electropotential, as well as decrease in pH, correlated with resumption of cellular activity on transfer into a defined medium. Eight amino acids were found to be essential for growth, while the remaining ten amino acids fell into four categories according to the degree of independence the organism had with respect to them. The form of the B-6 vitamin present in the medium had an important influence on the synthesis of several amino acids. The synthesis and degradation of amino acids was investigated further. The enzyme systems involved in these functions could be described as three major groups: those enzymes involved in synthesis of amino acids and amino compounds from glucose, the enzymes responsible for the interconversion of threonine, glycine and serine, and those involved with glutamic acid, proline and arginine metabolism. Uptake of amino acids by both resting and growing cell suspensions was found to be by an efficient, non-energy dependent diffusion mechanism. During growth approximately 5% of the exogenously supplied glucose was assimilated into cell material, primarily as cell wall or nucleic acid components. Molar growth yields from glucose were found to be significantly greater than the theoretical value. The calculated ratio of reduced to oxidized coenzymes indicated a characteristic trend over the logarithmic and stationary growth phases. The relationship between amino acid metabolism, glucose dissimilation and assimilation, and overall metabolite and energy balances has been discussed with a view to understanding the extent of control and influence C. perfringens has on its own metabolic functions and on its environment. / Science, Faculty of / Microbiology and Immunology, Department of / Graduate

The acidities of 37 amino-substituted heterocyclic compounds have been determined using measurements of degree of ionization in DMSO/ water/0.011 M hydroxide ion and recently developed extrapolative procedures which are based on the Bunnett-Olseh (B.O.) and the Marziano-Cimino-Passerini (M.C.P.) methods. The compounds studied include 19 aminopyridines and pyrimidines and 18 derivatives of the biologically important nucleotide bases, cytosine, adenine and guanine. The UV/vis spectra of the neutral and anionic forms of the amino-pyridines and pyrimidines are well separated so that in evaluating the ionization ratios, changes of the spectra with medium composition were accounted for by measuring at the wavelength of maximum anionic absorption in each solution and by assuming that eA- and eHA vary linearly with HN . For the majority of the nucleotide derivatives, the neutral and anionic spectra were closely overlapping although different in shape. The ionization ratios were best evaluated by using the area under the spectral curve; spectral changes with solvent composition were accounted for by an adaptation of the method described above. Ionization ratios evaluated by the area method agreed well with those calculated by other methods as long as the anionic area was at least one and one-half times as great as the neutral area (or vice versa). The aminopyrimidines and purines define a new acidity function Hp . This function covers the range from water to 87.6 mole % DMSO and was established using 15 aminopyrimidines and purines. When Hp plotted against mole % DMSO, it rises less steeply than HN. This has been attributed to less extensive derealization of charge in the aminopyrimidine anion relative to that of an aniline (usually a nitroaniline) ionizing at the same solvent composition. The B.O. pKHA values of unsubstituted 2- and 4-aminopyridine, 2- and 4-aminopyrimidine and 2-amino-s-triazine, ranging from 23.50 to 14.91 are well-correlated by a Hammett plot using a values recently established for aza substituents and assuming the additivity of aza substituent effects, p. 4.99. The B.O. pKHA values of five substituted anilines, ranging from 25.50 to 23.13, fall close to this plot. All ten compounds are accommodated by a straight line, p value 5.10, which suggests that the aminoheterocycles may be considered to be aza-substituted anilines. It appears that the sensitivity of aniline and diphenylamine acidity to substituent effects is greater than previously believed. The findings in this work suggest that transmission of substituent effects through a heterocyclic and a benzene nucleus are equal and that an aza group may be regarded as a normal ring substituent. From the acidities of substituted aminopyridines and pyrimidines it appears that, to a first approximation, aza groups do not perturb the effects of other substituents on the same nucleus, although there may be a small resonance interaction between a +R group, such as chloro, and an aza group lying para to it. The same conclusion was reached from examination of the basicities of these compounds. / Science, Faculty of / Chemistry, Department of / Graduate

One common method of improving the nutritional quality of certain food proteins is through fortification with necessary amounts of limiting essential amino acids. This simple and convenient method, however, is not the best. Several disadvantages are associated with the addition of free amino acids to food proteins, such as changes in flavor and color, losses of added amino acids during food processing or cooking, differences in stability and metabolism between free amino acids and amino acids in proteins. Covalent attachment of the limiting amino acids, however, should eliminate these problems, and moreover, could improve the nutritional and functional properties of food proteins. In this study, therefore, an attempt to improve the nutritional value of the soy protein was made by using the carbodiimide condensation reaction to covalently bind methionine and tryptophan to soy protein. In order to confine as much as possible the binding, of amino acid to the protein a-carboxyl groups the soy protein isolate (SPI) was partially hydrolyzed with pepsin to increase the number of a-carboxyls in soy protein. Various conditions of the carbodiimide reaction were analysed by a fractional factorial design in an attempt to determine the factors affecting the amino acid binding to soy protein hydrolysate (SPH). Of the factors investigated, pH, SPH concentration, carbodiimide concentration, activation time and reaction time were found to significantly affect the methionine binding efficiency, whereas pH, SPH concentration, carbodiimide concentration, amino acid concentration and reaction temperature were found to significantly influence the tryptophan binding efficiency to SPH. To determine the best level for each of the selected factors an optimization of the carbodiimide reaction conditions was conducted by carrying out another factorial experiment. Thus, under the best condition found, the methionine and tryptophan contents of methionine - and tryptophan-bound SPH samples were increased 7.7-fold and 18.0-fold, respectively. An in vitro pepsin-pancreatin digestion test demonstrated that the bound amino acids were readily released. In order to improve the low yield of the final product, another analysis of the carbodiimide reaction conditions was carried out. Since the yield could not be markedly improved by this factorial design in which peptic SPH was used, SPI without preliminary hydrolysis was used as the starting material. A product with 95-99% yield was obtained and its methionine or tryptophan content was increased 6.3-fold or 11.3-fold, respectively. High digestibility was still maintained for these products. Gel filtration chromatography demonstrated that the carbodiimide reaction caused an increase in the molecular weights of soy protein fractions. Furthermore, gel filtration chromatography revealed that, there was no selective amino acid binding among the different soy protein fractions during the carbodiimide reaction. / Land and Food Systems, Faculty of / Graduate

In the origin of life on the primitive earth a major step must have involved the condensation of amino acids to form the first polypeptides. Several suggestions as to how this might have occurred have been made by other workers. One of the more appealing proposals is that the polymerization was catalyzed by clay minerals. It has been reported, for example, that L-apartic acid polymerizes significantly faster than D-aspartic in the presence of kaolinite in aqueous suspension at 90˚. In this work an attempt was made to repeat this report and extend the pH range to include values presumably present on the prebiotic earth. No evidence for polymerization of L-aspartic acid were found. Polymerization of glycine under dry conditions in the presence of kaolinite and sodium borate was also investigated. Although small amounts of glycylglycine and glycylglycylglycine were detected there was no evidence that the reaction is enhanced by the presence of kaolinite.

A dissertation submitted to the Faculty of Health Sciences, Umversity of the Witwatersrand, Johannesburg, for the degree of Masters of Science in Medicine (Virology). Johannesburg, 2001 / This dissertation examines HIV-1 subtype C nucleotide and amino acid Nef sequences from South African recently-infected (n=12) and chronically-infected individuals (n=9) and from AIDS patients (n=ll). The overall aim is to determine which regions of subtype C Nef are variable and which are conserved and to associate these regions with targeting by the immune system. Phylogenetic analysis and sequence alignments showed that there was no association of Nef variations with stage of disease. Nef amino acid alignments of 32 sequences showed a high degree of conservation in areas of functional and structural importance. The mean intrasubtype variation from these isolates was 15.8%, with sequences from the AIDS cohort being significantly (p<0.05) more variable than those from recent or chronic infection / IT2019

No description available.