Isolation, characterization and molecular cloning of a lipolytic enzyme secreted from Malassezia pachydermatis.
Lipophilic Malassezia species may induce catheter-associated sepsis in premature neonates and immunocompromised patients receiving parenteral lipid emulsions. To assess the participation of lipolytic enzymes in the pathogenesis of this yeast, we cloned a gene encoding the enzyme. A lipolytic enzyme in the culture supernatant of Malassezia pachydermatis was purified 210-fold to homogeneity. The enzyme showed high esterase activity toward p-nitrophenyl octanoate. The cDNA encoding the enzyme was cloned using a degenerate oligonucleotide primer constructed from the N-terminal amino acid sequence. The cDNA consisted of 1582 bp, including an open reading frame encoding 470 amino acids. The first 19 amino acids and the following 13 amino-acid sequence were predicted to be the signal peptides for secretion and prosequence, respectively. The predicted molecular mass of the 438-amino acid mature protein was 48 kDa. Analysis of the deduced amino acid sequence revealed that it contains the consensus motif (Gly-X-Ser-X-Gly), which is conserved among lipolytic enzymes. Homology investigations showed that the enzyme has similarities principally with 11 lipases produced by Candida albicans (29-34% identity) and some other yeast lipases.
Shibata
Nobuyuki
N
Second Department of Hygienic Chemistry, Tohoku Pharmaceutical University, Sendai, Miyagi, Japan. nshibata@tohoku-pharm.ac.jp
Okanuma
Noriko
N
Hirai
Kanako
K
Arikawa
Kumiko
K
Kimura
Mayumi
M
Okawa
Yoshio
Y
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002712:Chlorides / Q000494:pharmacology; D003001:Cloning, Molecular / Q000379:methods; D017931:DNA Primers / Q000737:chemistry; D018076:DNA, Complementary / Q000737:chemistry; D004271:DNA, Fungal / Q000737:chemistry; D004492:Edetic Acid / Q000494:pharmacology; D004952:Esters / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000737:chemistry* / Q000145:classification / Q000187:drug effects / Q000302:isolation & purification*; D008290:Malassezia / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity*; D008969:Molecular Sequence Data; D009181:Mycoses / Q000201:enzymology* / Q000382:microbiology; D010802:Phylogeny; D010830:Physostigmine / Q000494:pharmacology; D016415:Sequence Alignment; D014105:Tosyl Compounds / Q000494:pharmacology; D014774:Virulence
D002712:Chlorides; D017931:DNA Primers; D018076:DNA, Complementary; D004271:DNA, Fungal; D004952:Esters; D014105:Tosyl Compounds; C003614:4-toluenesulfonyl fluoride; D004492:Edetic Acid; D010830:Physostigmine; D008049:Lipase