Lysophospholipase I identified as a ghrelin deacylation enzyme in rat stomach.
Ghrelin, discovered in rat stomach as an endogenous growth hormone secretagogue, is octanoylated at the Ser3 residue. Since this octanoylation is essential for the functions of ghrelin, the enzymes that catalyze acylation for ghrelin biosynthesis and deacylation (deactivation step) must be considered as important regulators. We found that rat stomach homogenate contained ghrelin deacylation activity, and we isolated the active fractions by column chromatography. After sequencing and expressing candidate proteins, the ghrelin deacylation enzyme in the stomach was identified as lysophospholipase I (LysoPLA I). The enzyme properties were examined using recombinant rat LysoPLA I expressed in Escherichia coli. K(m) and V(max) values were determined as 6.5 microM and 2.3 micromol/min/mg for ghrelin and 2.2 x 10(2) microM and 0.5 micromol/min/mg for lysophosphatidylcholine (LysoPC), respectively. The deacylation of both substrates was inhibited by methyl arachidonyl fluorophosphonate (MAFP), which is known as an irreversible inhibitor of LysoPLA I. These results reveal that LysoPLA I catalyzes the removal of n-octanoic acid from ghrelin to form des-acyl ghrelin. Identification of the ghrelin deacylation enzyme in the stomach and a deacylation inhibitor will be helpful in investigating ghrelin biosynthesis.
Shanado
Yuka
Y
Tsukuba Research Institute, Novartis Institutes for BioMedical Research, Tsukuba, Japan.
Kometani
Motohiko
M
Uchiyama
Hidefumi
H
Koizumi
Shinichi
S
Teno
Naoki
N
D000215:Acylation; D000595:Amino Acid Sequence; D000818:Animals; D004789:Enzyme Activation; D005766:Gastrointestinal Contents / Q000737:chemistry* / Q000201:enzymology*; D054439:Ghrelin; D008969:Molecular Sequence Data; D008970:Molecular Weight; D036361:Peptide Hormones / Q000737:chemistry* / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013270:Stomach / Q000201:enzymology*; D013379:Substrate Specificity; D013869:Thiolester Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D054439:Ghrelin; D036361:Peptide Hormones; D011994:Recombinant Proteins; C402226:LYPLA1 protein, human; D013869:Thiolester Hydrolases