Cloning and expression of rabbit pancreatic phospholipase A2.
Pancreatic phospholipase A2 (PLA2; E.C. 3.1.1.4) has been cloned from a gt11 library made from poly A+ RNA of adult rabbit pancreas. PLA2 catalyzes the hydrolysis of the 2-acyl ester bond of 3-sn-phosphoglycerides. As the rabbits are classically used for the study of diet induced changes in lipid metabolism, as a prelude to studying the diet and age dependent changes in this enzyme, we have undertaken to clone it from a rabbit pancreatic library. Three full length clones were obtained from the rabbit pancreatic library when probed with a synthetic oligonucleotide derived from the conserved portion of the molecule. One of these clones is completely sequenced and analyzed. The sequence consists of 606 nucleotides with an open reading frame of 441 nucleotides, coding for 147 amino acids which include a 15 amino acid leader peptide and a seven amino acid propeptide. Northern blot analysis revealed a major mRNA band at 600bp. When compared to phospholipases A2 of other species, rabbit PLA2 exhibited considerable conservation both at nucleotide and the protein level. In vitro translation of synthetic mRNA obtained from T7 polymerase transcription of the cDNA yielded a protein of apparent molecular weight of 15kd similar to that predicted from its primary structure.
Kumar
V B
VB
Geriatric Research Education and Clinical Center, VA Medical Center, St. Louis, MO 63125.
D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA; D006801:Humans; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010741:Phospholipases A / Q000235:genetics*; D054467:Phospholipases A2; D012333:RNA, Messenger / Q000235:genetics; D011817:Rabbits; D017386:Sequence Homology, Amino Acid
D012333:RNA, Messenger; D004247:DNA; D010741:Phospholipases A; D054467:Phospholipases A2