Dimerization of Thermomonospora fusca beta-1,4-endoglucanase E2.
Unboiled Thermomonospora fusca endoglucanase E2 electrophoresed on SDS-polyacrylamide gels migrated in the range of 80-90 kDa, but when boiled it migrated in the 40-42-kDa range. Sedimentation equilibrium centrifugation as well as chemical cross-linking experiments confirmed that E2 is a dimer. The dimer was reversibly dissociated at low pH. The E2 dimer was stable up to 70 degrees C, but began to dissociate at this temperature after a 30-60-min incubation. A nondimerizing mutant was obtained using region-specific chemical mutagenesis. DNA sequencing of this mutant revealed a single base change that substituted Gly for Glu-263. Chemical modification of carboxylic acid residues in E2 disrupted the dimer interaction.
McGinnis
K
K
Section of Biochemistry, Molecular, and Cell Biology, Cornell University, Ithaca, New York 14853.
Kroupis
C
C
Wilson
D B
DB
D000192:Actinomycetales / Q000201:enzymology*; D015153:Blotting, Western; D002480:Cellulase / Q000737:chemistry*; D003432:Cross-Linking Reagents; D004220:Disulfides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005976:Glutaral; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D008970:Molecular Weight; D009154:Mutation; D010084:Oxidation-Reduction; D014461:Ultracentrifugation
D003432:Cross-Linking Reagents; D004220:Disulfides; D002480:Cellulase; D005976:Glutaral