PMID:7490753
| Citation |
Kimura, M and Ishihama, A (1995) Functional map of the alpha subunit of Escherichia coli RNA polymerase: amino acid substitution within the amino-terminal assembly domain. J. Mol. Biol. 254:342-9 |
|---|---|
| Abstract |
The alpha subunit of Escherichia coli RNA polymerase plays a key role in assembly of the core enzyme. In previous studies the amino-terminal domain consisting of 215 amino acid residues between positions 21 and 235 was identified to be involved in this assembly, and the sites for beta and beta' association were suggested to be located within or near the two conserved regions in this amino-terminal assembly domain of alpha. For detailed functional mapping, Ala was substituted for 26 highly conserved amino acids around residues 40, 80 and 170 to 210. The alpha-point mutants were analyzed in vitro for their abilities to form dimers and to assemble beta beta' subunits. New types of assembly-deficient mutants were identified: alpha-R45A (having substituted Ala for Arg at residue 45) dimerized but did not assemble beta (and beta') subunits; and alpha-L48A showed a decreased level of alpha 2 beta subassembly formation, indicating that this region (residues 45 to 48) is responsible for beta-binding. Isolation of two mutants, alpha-K86A and alpha-V173A, both forming alpha 2 beta but not alpha 2 beta beta' complex, confirmed our previous conclusion that two separated regions participate in beta'-binding. |
| Links |
PubMed Online version:10.1006/jmbi.1995.0621 |
| Keywords |
Amino Acid Sequence; DNA Mutational Analysis; DNA-Directed RNA Polymerases/chemistry; DNA-Directed RNA Polymerases/genetics; DNA-Directed RNA Polymerases/metabolism; Escherichia coli/enzymology; Escherichia coli/genetics; Molecular Sequence Data; Point Mutation; Protein Binding; Protein Conformation; Recombinant Proteins/metabolism; Sequence Homology, Amino Acid |
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