Thermostabilization by replacement of specific residues with lysine in a Bacillus alkaline cellulase: building a structural model and implications ofnewly formed double intrahelical salt bridges

Citation
T. Ozawa et al., Thermostabilization by replacement of specific residues with lysine in a Bacillus alkaline cellulase: building a structural model and implications ofnewly formed double intrahelical salt bridges, PROTEIN ENG, 14(7), 2001, pp. 501-504
Citations number
43
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Biochemistry & Biophysics
Journal title
PROTEIN ENGINEERING
ISSN journal
0269-2139 → ACNP
Volume
14
Issue
7
Year of publication
2001
Pages
501 - 504
Database
ISI
SICI code
0269-2139(200107)14:7<501:TBROSR>2.0.ZU;2-I
Abstract
An alkaline, mesophilic endo-1,4-beta -glucanase from alkaliphilic Bacillus sp. strain KSM-64 was significantly thermostabilized by replacement of bot h Asn179 and Asp194 with lysine by site-directed mutagenesis. Structural re modeling of the mutant enzyme newly generated by the double mutation sugges ted that Glu175-->Lys179 and Glu190-->Lys194 were the most plausible ion pa irs, both of which involved side chains at the i and i + 4 positions on the alpha (4)-helix from Glu175 to Ser195. By molecular dynamics simulations, the N-zeta hydrogens of Lys179 and Lys194 were found to coordinate with the carbonyl O-epsilon1 and O-epsilon2 of Glu175 and the carbonyl O-epsilon1 o f Glu190, respectively, with distances of around 2 Angstrom for all. These results confirm that the formation of these double intrahelical ion pairs ( salt bridges) is responsible for the thermostabilization by the double muta tion.