E. coli F1 ‐ATPase: Site‐directed mutagenesis of the β‐subunit
- 9 May 1988
- journal article
- Published by Wiley in FEBS Letters
- Vol. 232 (1) , 111-114
- https://doi.org/10.1016/0014-5793(88)80397-1
Abstract
Residues βGlu-181 and βGlu-192 of E. coli F1-ATPase (the DCCD-reactive residues) were mutated to Gln. Purified βGln-181 F1 showed 7-fold impairment of ‘unisite’ Pi formation from ATP and a large decrease in affinity for ATP. Thus the β-181 carboxyl group in normal F1 significantly contributes to catalytic site properties. Also, positive catalytic site cooperativity was attenuated from 5 × 104- to 548-fold in βGln-181 F1. In contrast, purified βGln-192 F1 showed only 6-fold reduction in ‘multisite’ ATPase activity. Residues βGly-149 and βGly-154 were mutated to Ile singly and in combination. These mutations, affecting residues which are strongly conserved in nucleotide-binding proteins, were chosen to hinder conformational motion in a putative ‘flexible loop’ in β-subunit. Impairment of purified F1-ATPase ranged from 5 to 61%, with the double mutant F1 less impaired than either single mutant. F1 preparations containing βIle-154 showed 2-fold activation after release from membranes, suggesting association with F0 restrained turnover on F1 in these mutantsKeywords
This publication has 26 references indexed in Scilit:
- Electrostatic radius of the gramicidin channel determined from voltage dependence of H+ ion conductanceBiophysical Journal, 1988
- Invitro mutated β subunits from the F1-ATPase of the thermophilic bacterium, PS3, containing glutamine in place of glutamic acid in positions 190 or 201 assembles with the α and γ subunits to produce inactive complexesBiochemical and Biophysical Research Communications, 1987
- ß‐Subunit of Escherichia coli F1‐ATPaseFEBS Letters, 1987
- Structure of the nucleotide‐binding domain in the β‐subunit of Escherichia coli F1‐ATPaseFEBS Letters, 1986
- Primary structure and subunit stoichiometry of F1-ATPase from bovine mitochondriaJournal of Molecular Biology, 1985
- Effect of dicyclohexylcarbodiimide on unisite and multisite catalytic activities of the adenosine triphosphatase of Escherichia coliBiochemistry, 1985
- Conformational flexibility and its functional significance in some protein moleculesTrends in Biochemical Sciences, 1979
- Substrate positions and induced-fit in crystalline adenylate kinaseJournal of Molecular Biology, 1977
- Two conformations of crystalline adenylate kinaseJournal of Molecular Biology, 1977
- Structural invariants in protein foldingNature, 1975