title: Essential Role for an M17 Leucine Aminopeptidase in Encystation of Acanthamoeba castellanii.
abstract: Encystation of Acanthamoeba leads to the formation of resilient cysts from vegetative trophozoites. This process is essential for parasite survival under unfavorable conditions such as starvation, low temperatures, and exposure to biocides. During encystation, a massive turnover of intracellular components occurs, and a large number of organelles and proteins are degraded by proteases. Previous studies with specific protease inhibitors have shown that cysteine and serine proteases are involved in encystation of Acanthamoeba, but little is known about the role of metalloproteases in this process. Here, we have biochemically characterized an M17 leucine aminopeptidase of Acanthamoeba castellanii (AcLAP) and analyzed its functional involvement in encystation of the parasite. Recombinant AcLAP shared biochemical properties such as optimal pH, requirement of divalent metal ions for activity, substrate specificity for Leu, and inhibition profile by aminopeptidase inhibitors and metal chelators with other characterized M17 family LAPs. AcLAP was highly expressed at a late stage of encystation and mainly localized in the cytoplasm of A. castellanii. Knockdown of AcLAP using small interfering RNA induced a decrease of LAP activity during encystation, a reduction of mature cyst formation, and the formation of abnormal cyst walls. In summary, these results indicate that AcLAP is a typical M17 family enzyme that plays an essential role during encystation of Acanthamoeba.
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Yu-Ran
authors.initials: YR
authors.lastname: Lee
authors.affiliation: Department of Parasitology and Tropical Medicine, and Institute of Health Sciences, Gyeongsang National University School of Medicine, Jinju 660-751, Republic of Korea.
authors.forename: Byoung-Kuk
authors.initials: BK
authors.lastname: Na
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Eun-Kyung
authors.initials: EK
authors.lastname: Moon
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Su-Min
authors.initials: SM
authors.lastname: Song
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: So-Young
authors.initials: SY
authors.lastname: Joo
authors.affiliation: Department of Parasitology, Dong-A University, College of Medicine, Busan 602-714, Republic of Korea.
authors.forename: Hyun-Hee
authors.initials: HH
authors.lastname: Kong
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Youn-Kyoung
authors.initials: YK
authors.lastname: Goo
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Dong-Il
authors.initials: DI
authors.lastname: Chung
authors.affiliation: Department of Parasitology and Tropical Medicine, Kyungpook National University School of Medicine, Daegu 700-422, Republic of Korea.
authors.forename: Yeonchul
authors.initials: Y
authors.lastname: Hong
mesh_terms: D048248:Acanthamoeba castellanii / Q000145:classification / Q000502:physiology*; D000595:Amino Acid Sequence; D002614:Chelating Agents / Q000494:pharmacology; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000187:drug effects; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D007931:Leucyl Aminopeptidase / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D000069289:Parasite Encystment / Q000235:genetics; D010802:Phylogeny; D016415:Sequence Alignment
chemical_list: D002614:Chelating Agents; D007931:Leucyl Aminopeptidase