Linked Life Data

12074698

Source:http://linkedlifedata.com/resource/pubmed/id/12074698

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
Pt 3
pubmed:dateCreated
2002-6-20
pubmed:abstractText
A 1.6 kb DNA fragment including the lftM gene, encoding a levan fructotransferase (LFTase) of Microbacterium sp. AL-210, was subcloned into a high-expression vector, pET-29b, and the recombinant enzyme was overexpressed in Escherichia coli. Most of the LFTase activity was detected in the cytoplasmic fraction after induction with isopropyl beta-d-thiogalactoside. The recombinant enzyme with a tag of six histidine residues at the C-terminus was purified 132-fold by affinity and gel-filtration chromatography. Analysis of the N-terminal amino acid sequence revealed that the first 42 amino acids were post-translationally cleaved off. The molecular mass of the purified LftM was approx. 54 kDa as determined by SDS/PAGE, which corresponded well with a predicted size from the nucleotide sequence of the lftM gene lacking 42 amino acids. The enzyme converted levan into difructose anhydride IV (DFA IV) with a K(m) of 2 mg/ml and a V(max) of 40.6 micromol/min at pH 7.0 and 40 degrees C. The pH-dependence study of the enzyme for DFA IV production showed that LftM had a broad pH optimum (5.0-8.0) and the pK(a) values obtained were 4.5 and 8.9 at 40 degrees C. These results suggest that the acidic residues at the active site may play important roles for the catalytic mechanism of the LFTase.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jun
pubmed:issn
1470-8744
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
35
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
199-203
pubmed:meshHeading
pubmed:year
2002
pubmed:articleTitle
Expression, purification and characterization of a recombinant levan fructotransferase.
pubmed:affiliation
Department of Microbiology, College of Natural Sciences, Pusan National University, Jangjeon-dong, Kumjung-ku, Busan, Korea.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't