title: Structure, molecular evolution, and hydrolytic specificities of largemouth bass pepsins.
abstract: The nucleotide sequences of largemouth bass pepsinogens (PG1, 2 and 3) were determined after molecular cloning of the respective cDNAs. Encoded PG1, 2 and 3 were classified as fish pepsinogens A1, A2 and C, respectively. Molecular evolutionary analyses show that vertebrate pepsinogens are classified into seven monophyletic groups, i.e. pepsinogens A, F, Y (prochymosins), C, B, and fish pepsinogens A and C. Regarding the primary structures, extensive deletion was obvious in S'1 loop residues in fish pepsin A as well as tetrapod pepsin Y. This deletion resulted in a decrease in hydrophobic residues in the S'1 site. Hydrolytic specificities of bass pepsins A1 and A2 were investigated with a pepsin substrate and its variants. Bass pepsins preferred both hydrophobic/aromatic residues and charged residues at the P'1 sites of substrates, showing the dual character of S'1 sites. Thermodynamic analyses of bass pepsin A2 showed that its activation Gibbs energy change (∆G(‡)) was lower than that of porcine pepsin A. Several sites of bass pepsin A2 moiety were found to be under positive selection, and most of them are located on the surface of the molecule, where they are involved in conformational flexibility. The broad S'1 specificity and flexible structure of bass pepsin A2 are thought to cause its high proteolytic activity.
authors.affiliation: Division of Biomolecular Science, Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan; Department of Health and Nutrition, Nagoya Bunri University, Aichi 492-8213, Japan. Electronic address: y.miura@nsc.nagoya-cu.ac.jp.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Miura
authors.affiliation: Division of Biomolecular Science, Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan. Electronic address: m.suzuki@nsc.nagoya-cu.ac.jp.
authors.forename: Mieko
authors.initials: M
authors.lastname: Suzuki-Matsubara
authors.affiliation: Department of Health and Nutrition, Nagoya Bunri University, Aichi 492-8213, Japan. Electronic address: kageyama.takashi@nagoya-bunri.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kageyama
authors.affiliation: Division of Biomolecular Science, Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan. Electronic address: moriyama@nsc.nagoya-cu.ac.jp.
authors.forename: Akihiko
authors.initials: A
authors.lastname: Moriyama
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001492:Bass / Q000145:classification / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D019143:Evolution, Molecular; D029941:Fish Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D006868:Hydrolysis; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010434:Pepsin A / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010435:Pepsinogens / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013552:Swine; D013816:Thermodynamics
chemical_list: D018076:DNA, Complementary; D029941:Fish Proteins; D010435:Pepsinogens; D020033:Protein Isoforms; D012333:RNA, Messenger; D010434:Pepsin A
keywords: High proteolytic activity; Largemouth bass; Molecular evolution; Pepsin; Thermodynamicity