Characterization of glk, a gene coding for glucose kinase of Corynebacterium glutamicum.
The glk gene from Corynebacterium glutamicum was isolated by complementation using Escherichia coli ZSC113 (ptsG ptsM glk). We sequenced a total of 3072 bp containing the 969-bp open reading frame encoding glucose kinase (Glk). The glk gene has a deduced molecular mass of 34.2 kDa and contains a typical ATP binding site. Comparison with protein sequences revealed homologies to Glk from Streptomyces coelicolor (43%) and Bacillus megaterium (35%). The glk gene in C. glutamicum was inactivated on the chromosome via single crossover homologous recombination and the resulting glk mutant was characterized. Interestingly, the C. glutamicum glk mutant showed poor growth on rich medium such as LB medium or brain heart infusion medium in the presence or absence of glucose, fructose, maltose or sucrose as the sole carbon source. Growth yield was reduced significantly when maltose was used as the sole carbon source using minimal medium. The growth defect of glk mutant on rich medium was complemented by a plasmid-encoded glk gene. A chromosomal glk-lacZ fusion was constructed and used to monitor glk expression, and it was found that glk was expressed constitutively under all tested conditions with different carbon sources.
Park
S Y
SY
Environmental Bioresources Laboratory, Korea Research Institute of Bioscience and Biotechnology, College of Engineering, Yonsei University, Seoul, Korea.
Kim
H K
HK
Yoo
S K
SK
Oh
T K
TK
Lee
J K
JK
D000595:Amino Acid Sequence; D001665:Binding Sites; D002241:Carbohydrates; D003001:Cloning, Molecular; D003352:Corynebacterium / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D003470:Culture Media; D018274:Electroporation; D004926:Escherichia coli; D005798:Genes, Bacterial; D005822:Genetic Vectors; D005941:Glucokinase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D007763:Lac Operon; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D016415:Sequence Alignment; D014170:Transformation, Genetic
D002241:Carbohydrates; D003470:Culture Media; D005941:Glucokinase