title: Cloning and expression of a chitinase gene from Eisenia fetida.
abstract: Chitin is the second most abundant biopolymer in nature and is an important resource. In this study, we identified a chitinase gene, named Eisenia fetida-Chitinase (EF-Chi) gene, of 1494 base pairs (bp) that encodes a protein of 498 amino acids as indicated by the corresponding mRNA sequence. The amino acid sequence of EF-Chi was similar to those of chitinases from Eisenia andrei (99%), Branchiostoma floridae (50%) and Oryzias latipes (49%), and a gene encoding mature EF-Chi was expressed in the GS115 strain of Pichia pastoris. The molecular mass of the purified recombinant EF-Chi (rEF-Chi) was estimated to be 60kDa and catalytically important residues of chitinases of the glycoside hydrolase (GH) family 18 were conserved in EF-Chi. The optimal catalytic temperature of rEF-Chi was identified as 60°C, and the hydrolytic product from colloidal chitin was N-acetyl-chitobiose, suggesting that EF-Chi is an exo-type enzyme.
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, 599-8531, Japan. Electronic address: mueda@biochem.osakafu-u.ac.jp.
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Ueda
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, 599-8531, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Shioyama
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, 599-8531, Japan.
authors.forename: Kei
authors.initials: K
authors.lastname: Nakadoi
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, 599-8531, Japan.
authors.forename: Masami
authors.initials: M
authors.lastname: Nakazawa
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, 599-8531, Japan.
authors.forename: Tatsuji
authors.initials: T
authors.lastname: Sakamoto
authors.affiliation: Core Research Facilities for Basic Science (Molecular Cell Biology), Jikei University School of Medicine, 3-25-8 Nishishinbashi, Minato-ku, Tokyo, 105-8461, Japan.
authors.forename: Takeo
authors.initials: T
authors.lastname: Iwamoto
authors.affiliation: Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1, Nasahara, Takatsuki, Osaka, 569-1094, Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Sakaguchi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002688:Chitinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003001:Cloning, Molecular; D004795:Enzyme Stability; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009835:Oligochaeta / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D010843:Pichia / Q000235:genetics; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D002688:Chitinases
keywords: Chitinase; Earthworm; Eisenia fetida; Glycoside hydrolase (GH) family 18