A histidine binding protein ofEscherichia coli: a component of cystine binding protein ofEscherichia coli

Summary Commercially obtained cystine binding protein (CBP), an osmotic shock protein ofEscherichia coli, was studied in an effort to determine its binding characteristics. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS/PAGE) analysis of commercially obtained CBP showed three protein bands. N-terminal amino acid microsequencing and subsequent computer search revealed that the sequence of one of these proteins (25-kDa) was nearly identical to histidine binding protein (HisJ) ofSalmonella typhimurium. Purification of CBP by HPLC yielded four protein peaks, of which one bound histidine exclusively. Binding was maximal at pH 5.0 to 6.0, at 4°C, did not require calcium or magnesium ions and was not inhibited by reduction of CBP disulfide bonds. Amino acids other than histidine or cystine did not bind to CBP. These data show that commercially available CBP is not a homogenous protein; it contains a histidine as well as a cystine binding component.     

Developing a computer control system for a flexible manufacturing cell

This article focuses on the development of a computer control system for a flexible manufacturing cell. The paper outlines the various modules that are required for a computer control system and elaborates on the development of some of these modules. This is part of an ongoing research project at the University of the Witwatersrand. The aim of the project is to demonstrate the viability of flexible manufacturing systems to South African industry. Thus the main constraints of the project were cost, ease of implementation, and ease of maintainability. This necessitated the use of existing machinery and of computer systems and software that were readily available and already familiar to industry.     

Isolation from haddock tissue of psychrophilic bacteria with maximum growth temperature below 20 degrees C.

Protoplast fusion was investigated as a technique for genetically manipulating two lignin-degrading Streptomyces strains, Streptomyces viridosporus T7A and Streptomyces setonii 75Vi2. Four of 19 recombinants tested showed enhanced production of acid-precipitable polymeric lignin (APPL), producing 155 to 264% more APPL from corn stover lignocellulose than was produced by the wild-type S. viridosporus T7A. APPLs are lignin degradation intermediates known to be potentially valuable chemical products produced by bioconversion of lignin with Streptomyces spp. The prospects of utilizing protoplast fusion to construct APPL-overproducing Streptomyces strains was considered especially promising.     

Use of monoclonal antibody probes against rat hepatic cytochromes P-450c and P-450d to detect immunochemically related isozymes in liver microsomes from different species

Nine distinct monoclonal antibodies raised against purified rat liver cytochrome P-450c react with six different epitopes on the antigen, and one of these epitopes is shared by cytochrome P-450d. None of these monoclonal antibodies recognize seven other purified rat liver isozymes (cytochromes P-450a, b, and e-i) or other proteins in the cytochrome P-450 region of "Western blots" of liver microsomes. Each of the monoclonal antibodies was used to probe "Western blots" of liver microsomes from untreated, or 3-methylcholanthrene-, or isosafrole-treated animals to determine if laboratory animals other than rats possess isozymes immunochemically related to cytochromes P-450c and P-450d. Two protein-staining bands immunorelated to cytochromes P-450c and P-450d were observed in all animals treated with 3-methylcholanthrene (rabbit, hamster, guinea pig, and C57BL/6J mouse) except the DBA/2J mouse, where no polypeptide immunorelated to cytochrome P-450c was detected. The conservation of the number of rat cytochrome P-450c epitopes among these species varied from as few as two (guinea pig) to as many as five epitopes (C57BL/6J mouse and rabbit). The relative mobility in sodium dodecyl sulfate-gels of polypeptides immunorelated to cytochromes P-450c and P-450d was similar in all species examined except the guinea pig, where the polypeptide related to cytochrome P-450c had a smaller Mr than cytochrome P-450d. With the use of both monoclonal and polyclonal antibodies, we were able to establish that purified rabbit cytochromes P-450 LM4 and P-450 LM6 are immunorelated to rat cytochromes P-450d and P-450c, respectively.     

Vibrational raman spectra of lipid systems containing amphotericin B

Resonance-enhanced and normal vibrational Raman spectra were observed for both multilamellar and single-wall vesicle assemblies of dimyristoyl phosphatidylcholine containing amphotericin B, a channel-forming polyene antibiotic, and cholesterol. The decrease in the frequency of the polyene antibiotic C  C stretching mode at 1556 cm^?1 and the increase in intensity of the CCH in-plane deformation mode at 1002 cm^?1 indicate that amphotericin B is ordered in a lipid-cholesterol medium similarly to the solid, but is surrounded by a slightly more polar environment. The intensity of the C  C stretching mode I₁₅₅₆ decreases 4-fold during the broadened gel to liquid crystalline phase transition (16–32°C) of dimyristoyl lecithin-cholesterol (4 : 1) multilayers. Other resonance-enhanced vibrations of amphotericin B exhibit similar behavior. For amphotericin B in pure dimyristoyl lecithin multilayer or vesicle systems, however, the vibrational intensity associated with the C  C stretching mode remains constant during the melting of lipid hydrocarbon chains. In addition, a third effect occurs in liquid crystalline egg lecithin-cholesterol (4 : 1, mol ratio) multilayers in which I₁₅₅₆ first increases by 25% between 3 and 25°C, in parallel with the loss of active channels, and then remains constant as the temperature increases from 25 to 42°C. This latter intensity pattern is masked in the dimyristoyl lecithin-cholesterol system by the overwhelming effect upon the C  C mode from changes in the lipid chain packing characteristics which occur during the phase transition.The broadened phase transition in 4 : 1 dimyristoyl lecithin-cholesterol multilayers (16–32°C), as followed by the ratio of intensities at 2880 and 2850 cm^?1 (asymmetric and symmetric methylene C-H stretching modes, respectively) is slightly narrowed by the addition of amphotericin B, and effect from which a binding stoichiometry at 24° of 1 : 1 amphotericin B : cholesterol is estimated. This stoichiometry was confirmed by differential calorimetric scans, which also show the presence of a peak proportional to cholesterol content.Raman I_2880/2850 peak height ratios in pure dimyristoyl lecithin bilayers were increased over the 14–38°C range by amphotericin B, a spectral effect which suggests an ordering of the lipid matrix perhaps as a consequence of the polyene binding to the bilayer surface. For bilayers containing cholesterol, the ratios of intensities of the 2935 cm^?1 feature, composed mainly of acyl chain terminal methyl and underlying methylene C-H stretching modes, to the 2850 cm^?1 feature are significantly increased by amphotericin B. This effect indicates that the antibiotic penetrates the bilayer in the lipid-sterol system.     

Effect of solution non-ideality on erythrocyte volume regulation

A non-ideal, hydrated, non-dilute pseudo-binary salt-protein-water solution model of the erythrocyte intracellular solution is presented to describe the osmotic behavior of human erythrocytes. Existing experimental activity data for salts and proteins in aqueous solutions are used to formulate van Laar type expressions for the solvent and solute activity coefficients. Reasonable estimates can therefore be made of the non-ideality of the erythrocyte intracellular solution over a wide range of osmolalities. Solution non-ideality is shown to affect significantly the degree of solute polarization within the erythrocyte intracellular solution during freezing. However, the non-ideality has very little effect upon the amount of water retained within erythrocytes cooled at sub-zero temperatures.