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A ‘Gram‐negative‐type’ DNA polymerase III is essential for replication of the linear chromosome of Streptomyces coelicolor A3(2)
Author(s) -
Flett Fiona,
De Mello JungmannCampello Diana,
Mersinias Vassilios,
Koh Susie L.M.,
Godden Robert,
Smith Colin P.
Publication year - 1999
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.1999.01237.x
Subject(s) - biology , streptomyces coelicolor , genetics , dna , dna replication , chromosome , polymerase , dna polymerase , streptomyces , computational biology , gene , bacteria
The Streptomyces coelicolor dnaE gene, encoding the catalytic α‐subunit of DNA polymerase III (pol III) was isolated by genetic complementation of a temperature‐sensitive DNA replication mutant, S. coelicolor ts‐38. The deduced protein sequence (1179 residues) is highly similar to the Escherichia coli ‐type pol III α‐subunit, rather than to the PolC‐type α‐subunit that is known to be essential for replication in the ‘low G + C’ Gram‐positive bacteria such as Bacillus subtilis The dnaE gene is able to restore replication to a ‘slow stop’ mutant (ts‐38) and a ‘fast stop’ mutant (ts‐114); the dnaE gene of ts‐38 carries a single amino acid substitution (Glu‐802 to Lys), and the mutation in ts‐114 has been mapped between codons 697 and 1062 of dnaE . Mutant ts‐38 is considered to be defective in assembly of the multisubunit pol III holoenzyme and, hence, in initiation of replication, whereas ts‐114 is defective in chain elongation. This study provides the first evidence that a DnaE‐type pol III is essential for replication in a Gram‐positive bacterium. In addition, the complementation studies suggest that the C‐terminal 117 residues are not essential for DnaE function in S. coelicolor . When integrated at a distant site on the chromosome, a fragment containing the 3′ half of dnaE (codons 697–1179) is capable of rescuing ts‐38 (but not ts‐114) at the restrictive temperature; it was demonstrated that homogenotization was responsible for this phenomenon.