title: Investigation of Serine-Proteinase-Catalyzed Peptide Splicing in Analogues of Sunflower Trypsin Inhibitor 1 (SFTI-1).
abstract: Serine-proteinase-catalyzed peptide splicing was demonstrated in analogues of the trypsin inhibitor SFTI-1: both single peptides and two-peptide chains (C- and N-terminal peptide chains linked by a disulfide bridge). In the second series, peptide splicing with catalytic amount of proteinase was observed only when formation of acyl-enzyme intermediate was preceded by hydrolysis of the substrate Lys-Ser peptide bond. Here we demonstrate that with an equimolar amount of the proteinase, splicing occurs in all the two-peptide-chain analogues. This conclusion was supported by high resolution crystal structures of selected analogues in complex with trypsin. We showed that the process followed a direct transpeptidation mechanism. Thus, the acyl-enzyme intermediate was formed and was immediately used for a new peptide bond formation; products associated with the hydrolysis of the acyl-enzyme were not observed. The peptide splicing was sequence- not structure-specific.
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Natalia
authors.initials: N
authors.lastname: Karna
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Anna
authors.initials: A
authors.lastname: Łęgowska
authors.affiliation: Department of Microbiology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland.|Malopolska Centre of Biotechnology, Jagiellonian Univeristy, Gronostajowa 7a, 30-387, Krakow, Poland.
authors.forename: Stanisław
authors.initials: S
authors.lastname: Malicki
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Dawid
authors.initials: D
authors.lastname: Dębowski
authors.affiliation: Department of Microbiology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland.
authors.forename: Przemysław
authors.initials: P
authors.lastname: Golik
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Agata
authors.initials: A
authors.lastname: Gitlin
authors.affiliation: Department of Microbiology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland.
authors.forename: Przemysław
authors.initials: P
authors.lastname: Grudnik
authors.affiliation: Department of Microbiology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland.
authors.forename: Benedykt
authors.initials: B
authors.lastname: Wladyka
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Brzozowski
authors.affiliation: Department of Microbiology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland.|Malopolska Centre of Biotechnology, Jagiellonian Univeristy, Gronostajowa 7a, 30-387, Krakow, Poland.
authors.forename: Grzegorz
authors.initials: G
authors.lastname: Dubin
authors.affiliation: Department of Bioorganic Chemistry, Faculty of Chemistry, University of Gdansk, Wita Stwosza 63, 80-308, Gdansk, Poland.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Rolka
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D018360:Crystallography, X-Ray; D006368:Helianthus / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D010456:Peptides, Cyclic / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*; D057057:Serine Proteases / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D014357:Trypsin / Q000737:chemistry / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology*
chemical_list: D010455:Peptides; D010456:Peptides, Cyclic; C448501:SFTI-1 peptide, sunflower; D014361:Trypsin Inhibitors; D057057:Serine Proteases; D014357:Trypsin
keywords: SFTI-1; enzymes; inhibitors; peptide splicing