Pepsinogen and pepsin from the stomach of smooth hound (Mustelus mustelus):: Purification, characterization and amino acid terminal sequences

被引:41
作者
Bougatef, Ali [1 ]
Balti, Raft [1 ]
Ben Zaied, Saida [1 ]
Souissi, Nabil [2 ]
Nasri, Moncef [1 ]
机构
[1] Ecole Natl Ingenieurs Sfax, Lab Genei Enzymatigue Microbiol, Sfax, Tunisia
[2] Inst Natl Sci & Technol Mer, Sfax, Tunisia
关键词
pepsinogen; pepsin; biochemical characterization; stomach; smooth hound (Mustelus mustelus);
D O I
10.1016/j.foodchem.2007.08.077
中图分类号
O69 [应用化学];
学科分类号
081704 ;
摘要
Pepsinogen from the stomach of smooth hound (Mustelus mustelus) was purified to homogeneity by 20-70% ammonium sulphate precipitation, Sephadex G-100 gel filtration and DEAE-cellulose anion exchange chromatography with a 9.4-fold increase in specific activity and 38.36% recovery. Upon activation at pH 2.0, M. mustelus pepsinogen was converted to active form in one-step pathway. Molecular weights of the purified pepsinogen and the active pepsin were estimated to be 40,000 and 35,000 Da using SDS-PAGE and gel filtration, respectively. The optimum pH and temperature for the pepsin activity were pH 2.0 and 40 degrees C, respectively, using haemoglobin as a substrate. Activity was completely inhibited by Pepstatin A but not by phenylmethylsulphonyl fluoride, a serine-protease inhibitor and ethylenediaminetetraacetic acid, a metalloenzyme inhibitor. The N-terminal amino acid sequences of the first 15 amino acids of the activation segment of the pepsinogen and the first 20 amino acids of the active pepsin were LLRVPLRKGKSTLDV and ATEPLSNYLDSSYFGDISIG, respectively. M. mustelus pepsinogen, which showed high homology to rat C pepsinogen, had Thr-Leu-Asp sequence at amino acid positions 12-14 not found in all pepsinogen sequences. A remarkable substitution was found in the activation segment of M. mustelus pepsinogen: the Arg-13 conserved in all gastric proteinases, whose sequences are known, is replaced by Leu-13. (C) 2007 Elsevier Ltd. All rights reserved.
引用
收藏
页码:777 / 784
页数:8
相关论文
共 47 条
[1]   The estimation of pepsin, trypsin, papain, and cathepsin with hemoglobin [J].
Anson, ML .
JOURNAL OF GENERAL PHYSIOLOGY, 1938, 22 (01) :79-89
[2]   ISOLATION AND CHARACTERIZATION OF PEPSIN FROM POLAR COD (BOREOGADUS-SAIDA) [J].
ARUNCHALAM, K ;
HAARD, NF .
COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY B-BIOCHEMISTRY & MOLECULAR BIOLOGY, 1985, 80 (03) :467-473
[3]   COVALENT STRUCTURE OF CHICKEN PEPSINOGEN [J].
BAUDYS, M ;
KOSTKA, V .
EUROPEAN JOURNAL OF BIOCHEMISTRY, 1983, 136 (01) :89-99
[4]  
BOVEY FA, 1960, ENZYMES, V4, P63
[5]  
BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3
[6]   Secretagogues and growth factors in fish and crustacean protein hydrolysates [J].
Cancre, I ;
Ravallec, R ;
Van Wormhoudt, A ;
Stenberg, E ;
Gildberg, A ;
Le Gal, Y .
MARINE BIOTECHNOLOGY, 1999, 1 (05) :489-494
[7]   CONVERSION OF PEPSINOGEN INTO PEPSIN IS NOT A ONE-STEP PROCESS [J].
DYKES, CW ;
KAY, J .
BIOCHEMICAL JOURNAL, 1976, 153 (01) :141-144
[8]  
FOLTMANN B, 1981, ESSAYS BIOCHEM, V17, P52
[9]  
Fruton J. S., 1971, ENZYMES, V3, P119, DOI DOI 10.1016/S1874-6047(08)60395-9
[10]   RECOVERY OF PROTEINASES AND PROTEIN HYDROLYSATES FROM FISH VISCERA [J].
GILDBERG, A .
BIORESOURCE TECHNOLOGY, 1992, 39 (03) :271-276