Detection and Characterization of Novel Metal-Binding Proteins: Third Year (Progress) Report 1986--1987.

摘要

The research proposed and carried out under this grant concerned the detection and characterization of novel metal-binding proteins. In particular, the research goals as outlined in the original proposal were to detect novel metalloproteins from unusual bacteria such as Thiobacillus and Sulfolobus, to characterize one or more such novel metal-binding proteins, and as a long range goal beyond the scope of the proposal, to explore the conditions necessary for an industrial or scientific application of metal-binding proteins and to determine its relative advantages and limitations for the proposed application. Naturally occurring, metal-binding proteins are generally not a major component of living organisms and are thus difficult to identify and isolate. An important part of this research involved the development of techniques that would enable us to examine the proteins of bacteria for the presence of novel metal-binding proteins. Since many metal-binding proteins are rich in cysteine, we placed emphasis on techniques that would detect cysteine in proteins. The techniques that we examined included polyacrylamide gel electrophoresis of ( sup 35 S)cysteine-labeled cellular proteins, fast protein liquid chromatography (FPLC), and high-pressure liquid chromatography (HPLC) with on-line detection of copper or sulfhydryl groups. We demonstrated that these techniques work in model systems (yeast metallothionein and synthetic metal-binding polypeptides), and used them to examine the cellular proteins of Acidiphilium organovorum. Unfortunately, we were unable to identify any novel metal-binding proteins in this unusual, metal-resistant bacterium. 10 refs., 5 figs., 2 tabs. (ERA citation 13:055068)