{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,9]],"date-time":"2026-01-09T03:29:12Z","timestamp":1767929352942,"version":"3.49.0"},"reference-count":26,"publisher":"Springer Science and Business Media LLC","issue":"1","content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["BMC Microbiol"],"published-print":{"date-parts":[[2013,12]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:sec><jats:title>Background<\/jats:title><jats:p>Leucine aminopeptidase (LAP) has been known to be a housekeeping protease, DNA-binding protein and repressor or activator in the operon regulation of virulence-associated genes in several bacterial species. LAP activity was consistently detected in overnight cultures of<jats:italic>Burkholderia pseudomallei,<\/jats:italic>the causative agent of melioidosis and this enzyme was partially purified and characterised in this study. The intra- and inter-species nucleotide and deduced amino acid sequence variation of LAP encoding gene (<jats:italic>pep<\/jats:italic>A) was determined. A<jats:italic>pep<\/jats:italic>A\/PCR-RFLP assay was designed to facilitate the identification of major LAP sequence types amongst clinical and environmental isolates of<jats:italic>B. pseudomallei<\/jats:italic>.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>LAP activity was detected in<jats:italic>B. pseudomallei<\/jats:italic>culture supernantants by zymographic analysis. Optimum activity was at pH\u00a09 and stable at 50\u00b0C. Enhanced enzymatic activity was observed in the presence of metallic ions Mg<jats:sup>2+<\/jats:sup>, Ca<jats:sup>2+<\/jats:sup>, Na<jats:sup>+<\/jats:sup>and K<jats:sup>+<\/jats:sup>. LAP activity was inhibited by EDTA, 1,10-phenanthroline, amastatin, Mn<jats:sup>2+<\/jats:sup>and Zn<jats:sup>2+<\/jats:sup>. Sequence analysis of the complete nucleotide and deduced amino acid sequences of LAP-encoding (<jats:italic>pep<\/jats:italic>A) gene showed close genetic relatedness to<jats:italic>B. mallei<\/jats:italic>(similarity 99.7%\/99.6%), but not with<jats:italic>B. thailandensis<\/jats:italic>(96.4%\/96.4%). Eight<jats:italic>pep<\/jats:italic>A sequence types were identified by comparison with a 596\u00a0bp DNA fragment encompassing central regions of the<jats:italic>pep<\/jats:italic>A gene. A<jats:italic>pep<\/jats:italic>A\/PCR-RFLP was designed to differentiate<jats:italic>pep<\/jats:italic>A sequence types. Based on restriction analysis with<jats:italic>Stu<\/jats:italic>I and<jats:italic>Hinc<\/jats:italic>II enzymes of the amplified<jats:italic>pep<\/jats:italic>A gene, clinical and environmental isolates showed different predominant RFLP types. Type I was the most predominant type amongst 73.6% (67\/91) of the clinical isolates, while Type II was predominant in 55.6% (5\/9) of the environmental isolates.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>This study showed that LAP is a secretory product of<jats:italic>B. pseudomallei<\/jats:italic>with features similar to LAP of other organisms. Identification of major LAP sequence types of<jats:italic>B. pseudomallei<\/jats:italic>was made possible based on RFLP analysis of the<jats:italic>pep<\/jats:italic>A gene. The high LAP activity detected in both<jats:italic>B. pseudomallei<\/jats:italic>and<jats:italic>B. thailandensis<\/jats:italic>, suggests that LAP is probably a housekeeping enzyme rather than a virulence determinant.<\/jats:p><\/jats:sec>","DOI":"10.1186\/1471-2180-13-110","type":"journal-article","created":{"date-parts":[[2013,5,17]],"date-time":"2013-05-17T18:14:29Z","timestamp":1368814469000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":15,"title":["Enzymatic and molecular characterisation of leucine aminopeptidase of Burkholderia pseudomallei"],"prefix":"10.1186","volume":"13","author":[{"given":"Siew Mun","family":"Liew","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sun Tee","family":"Tay","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Savithiri D","family":"Puthucheary","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2013,5,17]]},"reference":[{"issue":"1","key":"1994_CR1","first-page":"160","volume":"29","author":"SM Liew","year":"2012","unstructured":"Liew SM, Tay ST, Wongratanacheewin S, Puthucheary SD: Enzymatic profiling of clinical and environmental isolates of Burkholderia pseudomallei. 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