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Isolation and sequence analysis of the pmi gene encoding phosphomannose isomerase of Streptococcus mutans
1Department of Biochemistry, Tokyo Dental College, Chiba City, Japan.
Abstract:
A gene encoding a phosphomannose isomerase from Streptococcus mutans GS-5 was identified immediately downstream from the fructokinase gene, scrK. Nucleotide sequence analysis of this region revealed an open reading frame (ORF) specifying a putative protein of 316 amino acids. The gene cloned in Escherichia coli expressed strong phosphomannose isomerase activity. The deduced amino acid sequence of the pmi gene has no significant similarity with any of the previously reported phosphomannose isomerase enzymes. Insertional inactivation of the upstream gene, scrK, in S. mutans also drastically reduced phosphomannose isomerase activity and the ability of the organism to utilize mannose as a sole carbon source. These results suggest that the S. mutans pmi gene constitutes an operon with the scrK gene.
Insights
Streptococcus mutans possesses a novel phosphomannose isomerase (pmi) gene, distinct from known enzymes. This gene, likely part of an operon with the fructokinase (scrK) gene, is crucial for mannose utilization.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Streptococcus mutans is a significant oral bacterium.
- Understanding carbohydrate metabolism in S. mutans is key to its biology.
- Phosphomannose isomerase (pmi) is an enzyme involved in mannose metabolism.
Purpose of the Study:
- To identify and characterize the phosphomannose isomerase gene in Streptococcus mutans GS-5.
- To investigate the relationship between the pmi gene and the fructokinase (scrK) gene.
- To determine the role of this pmi gene in mannose utilization by S. mutans.
Main Methods:
- Gene identification via nucleotide sequencing.
- Gene cloning and expression in Escherichia coli.
- Enzyme activity assays.
- Insertional inactivation of the scrK gene in S. mutans.
Main Results:
- A novel phosphomannose isomerase gene (pmi) was identified downstream of the scrK gene in S. mutans.
- The cloned pmi gene expressed significant phosphomannose isomerase activity in E. coli.
- The deduced amino acid sequence showed no similarity to known pmi enzymes.
- Inactivation of scrK in S. mutans reduced pmi activity and mannose utilization.
Conclusions:
- The identified pmi gene in S. mutans encodes a novel phosphomannose isomerase.
- The S. mutans pmi gene appears to form an operon with the scrK gene.
- This operon is essential for the efficient utilization of mannose as a carbon source by S. mutans.