title: Mass spectrometric detection of tyrosine sulfation in human pancreatic trypsinogens, but not in tumor-associated trypsinogen.
abstract: Trypsinogen-1 and -2 are well-characterized enzymes that are expressed in the pancreas and also in several other tissues. Many cancers produce trypsinogen isoenzymes that differ from the pancreatic ones with respect to substrate specificity and isoelectric point. These tumor-associated trypsinogens play a pivotal role in cancer progression and metastasis. The differences between these and the pancreatic isoenzymes have been suggested to be caused by post-translational modification, either sulfation or phosphorylation of a tyrosine residue. We aimed to elucidate the cause of these differences. We isolated trypsinogens from pancreatic juice and conditioned medium from a colon carcinoma cell line. Intact proteins, and tryptic and chymotryptic peptides were characterized by electrospray ionization mass spectrometry. We also used immunoblotting with antibody against phosphotyrosine and N-terminal sequencing. The results show that pancreatic trypsinogen-1 and -2 are sulfated at Tyr154, whereas tumor-associated trypsinogen-2 is not. Detachment of a labile sulfogroup could be demonstrated by both in-source dissociation and low-energy collision-induced dissociation in a tandem mass spectrometer. Tyrosine sulfation is an ubiquitous protein modification occurring in the secretory pathway, but its significance is often underestimated due to difficulties in its analysis. Sulfation is an almost irreversible modification that is thought to regulate protein-protein interactions and the activity of proteolytic enzymes. We conclude that the previously known differences in charge, substrate specificity and inhibitor binding between pancreatic and tumor-associated trypsinogens are probably caused by sulfation of Tyr154 in pancreatic trypsinogens.
authors: Department of Clinical Chemistry, Helsinki University Central Hospital, Finland.
authors: Outi
authors: O
authors: Itkonen
authors: Jari
authors: J
authors: Helin
authors: Juhani
authors: J
authors: Saarinen
authors: Nisse
authors: N
authors: Kalkkinen
authors: Konstantin I
authors: KI
authors: Ivanov
authors: Ulf-Håkan
authors: UH
authors: Stenman
authors: Leena
authors: L
authors: Valmu
mesh_terms: D000478:Alkylation; D000595:Amino Acid Sequence; D045744:Cell Line, Tumor; D006801:Humans; D008969:Molecular Sequence Data; D009369:Neoplasms / Q000378:metabolism* / Q000473:pathology; D010179:Pancreas / Q000201:enzymology*; D010766:Phosphorylation; D021241:Spectrometry, Mass, Electrospray Ionization; D013431:Sulfates / Q000378:metabolism*; D014362:Trypsinogen / Q000737:chemistry / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D013431:Sulfates; D014443:Tyrosine; D014362:Trypsinogen