Comparative Adsorption Studies with Synthetic, Structural Stability and Charge Mutants of Bacteriophage T4 Lysozyme.

Research output: Chapter in Book/Report/Conference proceedingBook chapter

Abstract

We have purified wild type, three structural stability mutants and four charge mutants of bacteriophage T4 lysozyme from E. coli strains harboring desired expression vectors. Structural stability mutants were produced by substitution of the isoleucine at amino acid position three, yielding a set of proteins with stabilities ranging from 1.2 kcal/mol greater, to 2.8 kcal/mol less, than that of the wild type. Charge mutants were produced by replacement of positively charged lysine residues with glutamic acid, yielding a set of molecules with formal charges ranging from +5 to +9 units. Adsorption kinetic data, along with the dodecyltrimethylammonium bromide-mediated elutability of each protein, has been monitored with in situ ellipsometry at hydrophobic and hydrophilic silica surfaces. A simple mechanism that allows adsorbing protein to adopt one of two states, each associated with a different resistance to elution and a different interfacial area occupied per molecule, has been used to assist interpretation of the adsorption data. Conditions implicit in the model have been used to estimate the fraction of molecules present on the surface just prior to surfactant addition that had adopted the more resistant state, and this fraction has been observed to correlate positively with resistance to elution. For the stability mutants, these properties were clearly related to protein stability as well. Concerning the charge mutants, results have not been clearly explainable in terms of protein net charge, but rather in terms of the probable influence of the location of each substitution relative to other mobile, solvent-exposed, charged side chains of the molecule.

Details

Authors
Organisations
External organisations
  • Oregon State University
  • Lund University
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Food Engineering

Keywords

Original languageEnglish
Title of host publicationProteins at Interfaces II
EditorsThomas A. Horbett, John L. Brash
PublisherThe American Chemical Society (ACS)
Pages52-65
Number of pages14
Volume602
ISBN (Electronic)9780841215276
ISBN (Print)9780841233041
Publication statusPublished - 1995
Publication categoryResearch
Peer-reviewedYes

Publication series

NameACS Symposium Series
PublisherAmerican Chemical Society
Volume602