title: Toward the smallest active subdomain of a TIM-barrel fold: insights from a truncated α-amylase.
abstract: AmyTM is a truncated mutant of the α-amylase of Bacillus stearothermophilus US100. It has been derived from the wild type amylase gene via a reading frame shift, following a tandem duplication of the mutant primer, associated to an Adenine base deletion. AmyTM was composed of 720 nucleotides encoding 240 amino acid residues out of 549 of the wild type. The AmyTM protein was devoided of the three catalytic residues but still retains catalytic activity. It is Ca-independent maltotetraose producing amylase, optimally active at pH 6 and 60°C, under monomeric or multimeric forms. AmyTM is the smallest functional truncated TIM barrel. It contains the βαβα unit as the minimal subdomain associated to an enzymatic function. The enzymatic activity can, until now, be attributed to the presence of the whole domain B, in the structure of AmyTM. This mutant revealed, for the first time, the regeneration of a catalytic site after its abolition. This fact may be considered as the restoration of a primitive active site, which was lost in the course of evolution toward more stable domains.
authors: Laboratoire de Métabolites et de Biomolécules, Centre de Biotechnologie de Sfax, Université de Sfax, B.P. 1177, 3018 Sfax, Tunisia. mamdouh.benali@cbs.rnrt.tn
authors: Mamdouh
authors: M
authors: Ben Ali
authors: Mehdi
authors: M
authors: Ghram
authors: Houda
authors: H
authors: Hmani
authors: Bassem
authors: B
authors: Khemakhem
authors: Richard
authors: R
authors: Haser
authors: Samir
authors: S
authors: Bejar
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001411:Geobacillus stearothermophilus / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D017510:Protein Folding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D000516:alpha-Amylases / Q000737:chemistry* / Q000235:genetics
chemical_list: D000516:alpha-Amylases