The complete nucleotide sequence (4,826 bp) of the cryptic plasmid pGA1 from Corynebacterium glutamicum was determined. DNA sequence analysis revealed four putative coding regions (open reading frame A [ORFA], ORFA2, ORFB, and ORFC). ORFC was identified as a rep gene coding for an initiator of plasmid replication (Rep) according to the high level of homology of its deduced amino acid sequence with the Rep proteins of plasmids pSR1 (from C. glutamicum) and pNG2 (from Corynebacterium diphtheriae). This function was confirmed by deletion mapping of the minimal replicon of pGA1 (1.7 kb) which contains only ORFC. Deletion derivatives of pGA1 devoid of ORFA exhibited significant decreases in the copy number in C. glutamicum cells and displayed segregational instability. Introduction of ORFA in trans into the cells harboring these deletion plasmids dramatically increased their copy number and segregational stability. The ORFA gene product thus positively influences plasmid copy number. This is the first report on such activity associated with a nonintegrating bacterial plasmid. The related plasmids pGA1, pSR1, and pNG2 lacking significant homology with any other plasmid seem to be representatives of a new group of plasmids replicating in the rolling-circle mode.

• MeSH
• Bacterial Proteins chemistry   genetics   physiology   MeSH
• Corynebacterium genetics   MeSH
• DNA-Binding Proteins * MeSH
• DNA Helicases chemistry   genetics   MeSH
• Molecular Sequence Data MeSH
• Open Reading Frames MeSH
• Plasmids genetics   MeSH
• Promoter Regions, Genetic MeSH
• Gene Expression Regulation, Bacterial MeSH
• Replicon genetics   MeSH
• Amino Acid Sequence MeSH
• Base Sequence MeSH
• Sequence Alignment MeSH
• Trans-Activators chemistry   genetics   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bacterial Proteins MeSH
• DNA-Binding Proteins * MeSH
• DNA Helicases MeSH
• ORFA protein, Corynebacterium MeSH Browser
• replication initiator protein MeSH Browser
• Trans-Activators MeSH

Cell protein profiles of submerged cultures of Streptomyces aureofaciens cultivated in the absence or presence of 12 microM benzyl thiocyanate (BT) were analyzed by one-dimensional SDS polyacrylamide gel electrophoresis. Substantial increase in the intensity of the 13, 35, 37, 60, and 100 kDa protein bands was observed in cultures treated with BT. Similar increase in the 35, 37, and 60 kDa bands was found in a mutant blocked in the last chlortetracycline biosynthesis step. Effect of BT on the solid medium-grown cultures was also observed, with a more intensive substrate mycelium pigmentation and alteration in the spore size and shape as the most characteristic features. Earlier studies of BT effect involving those on the stimulation of chlortetracycline biosynthesis are summarized and a possible signal-transducing mechanism is discussed from the point of view of adaptation of S. aureofaciens to the uncoupling of oxidative phosphorylation.

• MeSH
• Bacterial Proteins metabolism   MeSH
• Microscopy, Electron MeSH
• Streptomyces aureofaciens drug effects   metabolism   ultrastructure   MeSH
• Tetracyclines biosynthesis   MeSH
• Thiocyanates pharmacology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bacterial Proteins MeSH
• benzyl thiocyanate MeSH Browser
• Tetracyclines MeSH
• Thiocyanates MeSH