Title

Guanidination Of Proteins

Date of Award

1981

Availability

Article

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Biochemistry and Molecular Biology

Abstract

Earlier studies have indicated the marked resistance of two Pronase endopeptidases to denaturation in high concentrations of urea or guanidine hydrochloride (Siegel, S., and Awad, W. M. Jr. {1973} J. Biol. Chemistry 248, 3233-3240). One component has only a single residue of lysine and the other has none. The consideration arose that lysine-containing peptide segments may be less stable than those containing arginine because of the fluctuations of the side groups of the former residue. The small (epsilon)-amino groups may not be able to sustain solvation of the hydrophobic arm in an aqueous medium. Arginine residues have shorter hydrophobic arms, larger hydrophilic groups, and higher pKa's and thus may be less motile than lysine. This hypothesis was tested by quanidination of seven globular proteins (bovine carbonic anhydrase, chymotrypsinogen, (alpha)-lactalbumin, serum albumin, and ribonucclease; hen egg lysozyme; and horse heart cytochrome c). Conversion of lysine residues to homoarginine was between 90 to 99%. Tritium-hydrogen isotope exchange revealed that all proteins except lysozyme demonstrated reduced out-exchange after quanidination. The results with lysozyme were not unexpected since only this protein has a high arginine-to-lysine ratio. These findings suggest that high arginine-to-lysine ratios contribute to protein stability.Further studies with each protein demonstrate a marked reduction in solubility after quanidination; measurements were done at various pH values and also as a function of ammonium sulfate concentration. Gel filtration analysis of guanidinated proteins revealed in all cases except (alpha)-lactalbumin that the monomeric protein was studied during hydrogen-isotope exchange studies, thus indicating that stabilization was independent of aggregation. These findings suggest the possibility that the changes in solubility are not independent of the changes in stability. In fact proteins may be most soluble when they show conformational flexibility; solubility may be reduced with tighter conformations because the latter stabilize surface bonds between like protein molecules.

Keywords

Chemistry, Biochemistry

Link to Full Text

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