"Adult only" esterase/phospholipase A of the small-intestine brush border membrane: isolation, identification of the catalytic site, and biosynthesis.
We have isolated and investigated an esterase/phospholipase A (EC 3.1.1) which is an intrinsic protein of the small-intestine brush border membrane of adult but not of preweaning rabbits. This enzyme had been referred to previously as AdRabB [Boll, W., Schmid-Chanda, T., Semenza, G., & Mantei, N. (1993) J. Biol. Chem. 268, 12901-12911]. Its amino acid sequence shows four extensive, homologous repeats between the signal sequence and a hydrophobic stretch in the C-terminal region. We have identified a serine residue (Ser400) in the (unexpectedly) single catalytic site and indicate that this esterase is likely to operate by way of a Ser-His-Asp/Glu triad. This esterase/phospholipase A is synthesized as a single polypeptide chain of 170-180 kDa, agreeing well with the size deduced from its cognate cDNA sequence [Boll, W., et al. (1993) J. Biol. Chem. 268, 12901-12911], and it undergoes (at least) N-glycosylation. Once it has reached the brush border membrane, it is subjected in vivo to two types of proteolytic processing, presumably by pancreatic protease(s): a split after Arg263, with loss of the first N-terminal repeat, and two or more cleavages much farther downstream but still located in the luminal bulk of the protein; these splits lead to multiple bands of approximately 140, 125, 100, and 90 kDa.
Wacker
H
H
Department of Biochemistry, Swiss Federal Institute of Technology, ETH-Zentrum, Zurich.
Keller
P
P
von Balthazar
A K
AK
Semenza
G
G
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D004950:Esterases / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000201:enzymology*; D007421:Intestine, Small; D007700:Kinetics; D008297:Male; D008871:Microvilli / Q000201:enzymology*; D008969:Molecular Sequence Data; D009924:Organ Culture Techniques; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010449:Peptide Mapping; D010741:Phospholipases A / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D014162:Transfection
D010446:Peptide Fragments; D011994:Recombinant Proteins; D004950:Esterases; D010741:Phospholipases A