Expression and functional analysis of rat P23, a gut hormone-inducible isoform of trypsin, reveals its resistance to proteinaceous trypsin inhibitors.
Rat P23 is an isoform of trypsin (ogens) synthesized by rat acinar cells. Expression of P23 is stimulated strongly by caerulein, an analogue of cholecystokinin (CCK). However, the physiological relevance of rat P23 in healthy and pathological conditions such as caerulein-induced pancreatitis is largely unknown. Using recombinant P23 trypsinogen and reconstitution analysis of zymogen autoactivation, unique inhibitor-resistance characteristics of P23 were elucidated. P23 cDNA was expressed in Escherichia coli periplasm, yielding recombinant P23 trypsinogen. Autoactivation of zymogen granule contents from caerulein-induced rat pancreas was also studied. Activation kinetics of P23 by enterokinase was similar to those of rat anionic trypsinogen, which is a major isoform of trypsinogen. Interestingly, rat pancreatic secretory trypsin inhibitor (PSTI), which protects against deleterious activation of trypsinogens in zymogen granules, failed to inhibit P23 trypsin even with four-fold molar excess, at which concentration it effectively inhibited rat anionic trypsin to almost 100%. P23 trypsin also showed marked resistance to proteinaceous trypsin inhibitors such as soybean trypsin inhibitor and aprotinin. P23 trypsin activated by enterokinase dramatically accelerated the cascade of autoactivation of anionic trypsinogen even in the presence of PSTI. Taken together with a previous observation that P23 is specifically upregulated 14-fold by 24-h caerulein infusion, these results suggest that elevated levels of P23 should be taken into consideration in the mechanism of trypsinogens within the pancreas in pathological conditions.
Fukuoka
Shin-Ichi
S
Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611, Japan. fukuoka@food2.food.kyoto-u.ac.jp
Nyaruhucha
Cornelio M
CM
D000595:Amino Acid Sequence; D000818:Animals; D002108:Ceruletide; D003001:Cloning, Molecular; D004765:Enteropeptidase / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000378:metabolism; D010179:Pancreas / Q000378:metabolism; D010195:Pancreatitis / Q000139:chemically induced / Q000378:metabolism; D020033:Protein Isoforms / Q000737:chemistry; D051381:Rats; D014361:Trypsin Inhibitors / Q000494:pharmacology; D014362:Trypsinogen / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
D020033:Protein Isoforms; D014361:Trypsin Inhibitors; D002108:Ceruletide; D014362:Trypsinogen; D004765:Enteropeptidase