Sequence of the Corynebacterium glutamicum pyruvate carboxylase gene.

TitleSequence of the Corynebacterium glutamicum pyruvate carboxylase gene.
Publication TypeJournal Article
Year of Publication1998
AuthorsKoffas, MA, Ramamoorthi, R, Pine, WA, Sinskey, AJ, Stephanopoulos, G
JournalAppl Microbiol Biotechnol
Date Published1998 Sep
KeywordsAmino Acid Sequence, Blotting, Western, Cloning, Molecular, Corynebacterium, Cosmids, DNA, Bacterial, Gene Library, Humans, Immunoblotting, Molecular Sequence Data, Polymerase Chain Reaction, Pyruvate Carboxylase, Sequence Analysis, DNA, Sequence Homology, Amino Acid

Pyruvate carboxylase is an important anaplerotic enzyme replenishing oxaloacetate consumed for biosynthesis during growth, or lysine and glutamic acid production in industrial fermentations. We used regions of homology from pyruvate carboxylase sequences of 12 different species (corresponding to the ATP- and pyruvate-binding sites), to design polymerase chain reaction (PCR) primers for amplifying a fragment of the pyruvate carboxylase (pc) gene from C. glutamicum genomic DNA. This 850-base-pair fragment was used to probe a C. glutamicum cosmid library and four candidate pc cosmids were identified. The fragment was sequenced and the sequence of the complete gene was obtained by several rounds of primer synthesis, PCR on one of the positive cosmids, and sequencing. The C. glutamicum pc sequence shows 64% homology with the pc gene of Mycobacterium tuberculosis and 44% homology with the human pc gene. Regions of ATP, pyruvate and biotin binding have also been identified.

Alternate JournalAppl Microbiol Biotechnol
Citation Key211
PubMed ID9802220