Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker

Kawashima, Satoshi; Ikehata, Hiroki; Tada, Chihiro; Ogino, Tomohiro; Kakizaki, Hiromi; Ikeda, Mana; Fukushima, Hideto; Matsumiya, Masahiro

doi:10.3390/md14010022

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Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker

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Department of Marine Science and Resources, College of Bioresource Sciences, Nihon University, Fujisawa, Kanagawa 252-0880, Japan

Author to whom correspondence should be addressed.

Academic Editor: Takao Ojima

Mar. Drugs 2016, 14(1), 22; https://doi.org/10.3390/md14010022

Received: 15 December 2015 / Revised: 9 January 2016 / Accepted: 13 January 2016 / Published: 20 January 2016

(This article belongs to the Special Issue Green Chemistry Approach to Marine Products)

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Abstract

Fish express two different chitinases, acidic fish chitinase-1 (AFCase-1) and acidic fish chitinase-2 (AFCase-2), in the stomach. AFCase-1 and AFCase-2 have different degradation patterns, as fish efficiently degrade chitin ingested as food. For a comparison with the enzymatic properties and the primary structures of chitinase isozymes obtained previously from the stomach of demersal fish, in this study, we purified chitinase isozymes from the stomach of Japanese sardine Sardinops melanostictus, a surface fish that feeds on plankton, characterized the properties of these isozymes, and cloned the cDNAs encoding chitinases. We also predicted 3D structure models using the primary structures of S. melanostictus stomach chitinases. Two chitinase isozymes, SmeChiA (45 kDa) and SmeChiB (56 kDa), were purified from the stomach of S. melanostictus. Moreover, two cDNAs, SmeChi-1 encoding SmeChiA, and SmeChi-2 encoding SmeChiB were cloned. The linker regions of the deduced amino acid sequences of SmeChi-1 and SmeChi-2 (SmeChi-1 and SmeChi-2) are the longest among the fish stomach chitinases. In the cleavage pattern groups toward short substrates and the phylogenetic tree analysis, SmeChi-1 and SmeChi-2 were classified into AFCase-1 and AFCase-2, respectively. SmeChi-1 and SmeChi-2 had catalytic domains that consisted of a TIM-barrel (β/α)₈–fold structure and a deep substrate-binding cleft. This is the first study showing the 3D structure models of fish stomach chitinases. View Full-Text

Keywords: fish stomach chitinase isozymes; cDNA cloning; Japanese sardine Sardinops melanostictus; fish-specific acidic fish chitinase; long linker

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Kawashima, S.; Ikehata, H.; Tada, C.; Ogino, T.; Kakizaki, H.; Ikeda, M.; Fukushima, H.; Matsumiya, M. Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker. Mar. Drugs 2016, 14, 22. https://doi.org/10.3390/md14010022

AMA Style

Kawashima S, Ikehata H, Tada C, Ogino T, Kakizaki H, Ikeda M, Fukushima H, Matsumiya M. Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker. Marine Drugs. 2016; 14(1):22. https://doi.org/10.3390/md14010022

Chicago/Turabian Style

Kawashima, Satoshi, Hiroki Ikehata, Chihiro Tada, Tomohiro Ogino, Hiromi Kakizaki, Mana Ikeda, Hideto Fukushima, and Masahiro Matsumiya. 2016. "Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker" Marine Drugs 14, no. 1: 22. https://doi.org/10.3390/md14010022

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Stomach Chitinase from Japanese Sardine Sardinops melanostictus: Purification, Characterization, and Molecular Cloning of Chitinase Isozymes with a Long Linker

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