Purification and characterization of guinea pig gastric phospholipase A2 of the pancreatic type.
Guinea pig gastric mucosa and juice contained exceptionally high phospholipase-A2 activity, whereas the activity in the pancreas was only minimal. Phospholipases A2 were purified to homogeneity from these three tissues. Structural evidence, including the sequence of the NH2-terminal 41 residues, the amino-acid composition and the molecular mass (13902 +/- 3 Da) determined accurately by mass spectrometry, showed that the gastric mucosa enzyme belongs to the pancreatic type. An unique feature of the sequence is the substitution of Phe for the hitherto invariant Tyr28 in the calcium-binding loop of pancreatic phospholipases A2. The affinity of the guinea pig enzyme for Ca2+ in the presence of substrate was, however, identical to that of the rat enzyme with Tyr28, suggesting the interaction of a phenolic hydroxyl group of the Tyr with its neighboring residues is not significantly linked to the binding of Ca2+. The NH2-terminal sequences and immunochemical properties of the enzymes purified from the gastric juice and pancreas were identical to those of the gastric mucosa enzyme. The distribution of cells immunoreactive with anti-(gastric PLA2) immunoglobulin in the stomach was quite similar to that of the chief cells. Unlike in pancreas of other animals, the prophospholipase A2 was not detectable in gastric mucosa or juice homogenates treated with diisopropyl fluorophosphate or in column effluents during purification under acidic conditions. An appreciable prophospholipase-A2-activating activity was not detectable in gastric mucosa extracts at low pH relevant to gastric juice, using rat prophospholipase A2 as substrate. This opposes the activation of secreted proenzyme in the gastric juice.
Tojo
H
H
Department of Molecular Physiological Chemistry, Osaka University Medical School, Japan.
Ying
Z
Z
Okamoto
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000494:pharmacology; D005750:Gastric Juice / Q000201:enzymology*; D005753:Gastric Mucosa / Q000201:enzymology*; D006168:Guinea Pigs; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D008969:Molecular Sequence Data; D010179:Pancreas / Q000201:enzymology*; D010741:Phospholipases A / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification*; D054467:Phospholipases A2; D051381:Rats
D010741:Phospholipases A; D054467:Phospholipases A2; D002118:Calcium