Activities | Families | Sequences | Fold types | References | Help
B6db references: 8074505

type Journal Article
authors Malumbres, M.; Mateos, L. M.; Lumbreras, M. A.; Guerrero, C.; Martin, J. F.
title Analysis and expression of the thrC gene of Brevibacterium lactofermentum and characterization of the encoded threonine synthase
journal Appl Environ Microbiol
sel selected
ui 8074505
year (1994)
volume 60
number 7
pages 2209-19
keywords Amino Acid Sequence
abstract The thrC gene of Brevibacterium lactofermentum was cloned by complementation of Escherichia coli thrC auxotrophs. The gene was located by deletion mapping and complementation analysis in a 2.9-kb Sau3AI-HindIII fragment of the genome. This fragment also complemented a B. lactofermentum UL1035 threonine auxotroph that was deficient in threonine synthase. A 1,892-bp DNA fragment of this region was sequenced; this fragment contained a 1,446-bp open reading frame that encoded a 481-amino-acid protein having a deduced M(r) of 52,807. The gene was expressed in E. coli, by using the phage T7 system, as a 53- kDa protein. The promoter region subcloned in promoter-probe plasmids was functional in E. coli. A Northern analysis revealed that the gene was expressed as a monocistronic 1,400-nucleotide transcript. The transcription start point of the thrC gene was located by S1 mapping 6 bp upstream from the translation initiation codon, which indicated that this promoter was one of the leaderless transcription-initiating sequences. The threonine synthase overexpressed in B. lactofermentum UL1035 was purified almost to homogeneity. The active form corresponded to a monomeric 52.8-kDa protein, as shown by sodium dodecyl sulfate- polyacrylamide gel electrophoresis. The purified enzyme required pyridoxal phosphate as its only cofactor to convert homoserine phosphate into threonine.
last changed 2009/06/26 10:21

B6db references