2Kurchatov Institute, 123182 Moscow, Russia; fax: +7 (499) 1961704
3Bach Institute of Biochemistry, Russian Academy of Sciences, 119071 Moscow, Russia; fax: +7 (495) 954-2732
4Moscow State Pedagogical University, 119991 Moscow, Russia
* To whom correspondence should be addressed.
Received April 7, 2015; Revision received June 2, 2015
Treatment of native psychrophilic oligopeptidase B from Serratia proteamaculans (PSP, 78 kDa) with chymotrypsin (soluble or immobilized on modified porous glass MPG-PA) in the presence of 50% glycerol leads to production of a truncated enzyme form (PSP-Chtr, ~66 kDa), which retains activity toward the low molecular weight substrate of PSP, BAPNA, but in contrast to PSP, is active toward the protein substrate azocasein. It has been shown by MALDI-TOF mass-spectrometry that PSP-Chtr lacks the N-terminal region of the molecule that envelops the catalytic domain of PSP and supposedly prevents hydrolysis of high molecular weight substrates. It has also been established that the lacking fragment corresponds to the N-terminal highest rank element of the informational structure of PSP. This finding confirms the usefulness of the method of informational structure analysis for protein engineering of enzymes. A similar treatment of PSP with immobilized trypsin also led to production of a stable truncated enzyme form (PSP-Tr, ~75 kDa) which lacked 22 C-terminal amino acid residues and completely lost enzymatic activity, presumably because of changes in the nearest environment of His652 of the catalytic triad.
KEY WORDS: oligopeptidase B, Serratia proteamaculans, restricted proteolysis, azocasein, MALDI-TOF mass-spectrometry, analysis of informational protein structure