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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:dateCreated |
1995-4-27
|
| pubmed:databankReference | |
| pubmed:abstractText |
Triosephosphate isomerase (EC 5.3.1.1) from Lactococcus lactis was purified to electrophoretic homogeneity. Approximately 3 mg purified enzyme (specific activity 3300 U mg-1) was obtained from 70 g (wet wt) cells. In solution, triosephosphate isomerase (pI 4.0-4.4) was observed to exist as a homodimer (M(r) 57,000) of noncovalently linked subunits. The sequence of the first 37 amino acid residues from the NH2-terminus were determined by step-wise Edman degradation. This sequence, and that of a region conserved in all known bacterial triosephosphate isomerases, was used to design oligonucleotide primers for the synthesis of a lactococcal tpi probe by PCR. The probe was used to isolate a molecular clone of tpi from a lambda GEM11 library of L. lactis LM0230 DNA. The nucleotide sequence of tpi predicted a protein of 252 amino acids with the same NH2-terminal sequence as that determined for the purified enzyme and a subunit M(r) of 26,802 after removal of the NH2-terminal methionine. Escherichia coli cells harbouring a plasmid containing tpi had 15-fold higher triosephosphate isomerase activity than isogenic plasmid-free cells, confirming the identity of the cloned gene. Northern analysis of L. lactis LM0230 RNA showed that a 900 base transcript hybridized with tpi. The 5' end of the transcript was determined by primer extension analysis to be a G located 65 bp upstream from the tpi start codon. These transcript analyses indicated that in L. lactis, tpi is expressed on a monocistronic transcript. Nucleotide sequencing indicated that the DNA adjacent to tpi did not encode another Embden-Meyerhoff-Parnas pathway enzyme. The location of tpi on the L. lactis DL11 chromosome map was determined to be between map coordinates 1.818 and 1.978.
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| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Jan
|
| pubmed:issn |
1350-0872
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| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:volume |
141 ( Pt 1)
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
229-38
|
| pubmed:dateRevised |
2006-11-15
|
| pubmed:meshHeading |
pubmed-meshheading:7534588-Amino Acid Sequence,
pubmed-meshheading:7534588-Base Sequence,
pubmed-meshheading:7534588-Blotting, Northern,
pubmed-meshheading:7534588-Consensus Sequence,
pubmed-meshheading:7534588-DNA, Bacterial,
pubmed-meshheading:7534588-DNA Primers,
pubmed-meshheading:7534588-Gene Library,
pubmed-meshheading:7534588-Genes, Bacterial,
pubmed-meshheading:7534588-Lactococcus lactis,
pubmed-meshheading:7534588-Molecular Sequence Data,
pubmed-meshheading:7534588-Plasmids,
pubmed-meshheading:7534588-Polymerase Chain Reaction,
pubmed-meshheading:7534588-RNA, Bacterial,
pubmed-meshheading:7534588-Restriction Mapping,
pubmed-meshheading:7534588-Sequence Homology, Amino Acid,
pubmed-meshheading:7534588-Species Specificity,
pubmed-meshheading:7534588-Transcription, Genetic,
pubmed-meshheading:7534588-Triose-Phosphate Isomerase
|
| pubmed:year |
1995
|
| pubmed:articleTitle |
The Lactococcus lactis triosephosphate isomerase gene, tpi, is monocistronic.
|
| pubmed:affiliation |
Russell Grimwade School of Biochemistry, University of Melbourne, Parkville, Victoria, Australia.
|
| pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, Non-U.S. Gov't
|