Temperature characteristics of peptidase in chironomid larvae, potential fish prey, at various pH values

Keywords: chironomid larvae; peptidase; temperature dependence; temperature coefficients; activation energy; pH


The temperature dependence of casein- and hemoglobinlytic peptidases functioning in the whole organism of chironomid larvae Chironomus plumosus, food objects of adult benthophages and juvenile fish of various ecological groups, was studied within the temperature range of 0–70 ºС at different рН values (3.0, 5.0 and 7.4). The method of mixed samples was used to determine the activity and characteristics of enzymes. Homogenates of previously crushed and carefully mixed dozens of larvae were used as enzymatically active preparations. Activity of peptidases was assayed by the increase in tyrosine concentration using the Folin-Ciocalteu reagent. It is shown that the activity of peptidases that function in the tissues of chironomid larvae depends to a considerable extent on temperature and рН, but the pH has a smaller effect on the activity and the temperature dependence of casein- and hemoglobin-lytic peptidases than temperature. The temperature optimum of the studied peptidases of chironomid larvae corresponds to 40 ºС. The Q10 values in the zone of vital temperatures are slightly changed. They are, as a rule, increased in the zone of 30–40 ºС, and are sharply decreased in the zone of high temperatures. The values of activation energy of the process of hydrolysis of casein and hemoglobin in the zone of low and high temperatures are different. The Еact values of the process of hydrolysis of casein and hemoglobin at a temperature not exceeding 20 ºС are usually below those in the zone of higher temperatures (except for hemoglobin-lytic peptidases at pH 5.0). The data obtained indicate a significant effect of pH not only on the activity, but also on the temperature characteristics of peptidases that function in the body of chironomid larvae. Differences in the characteristics of casein- and hemoglobin-lytic peptidases in chironomid larvae at different temperatures and pH can influence the digestion in benthophages and fry of all fish species.


Anson, M. (1938). The estimation of pepsin, trypsin, papain and cathepsin with haemoglobin Journal of General Physiology, 22, 79–83.

Aoki, H., Ahsan, M. N., & Watabe, S. (2003). Molecular cloning and characterization of cathepsin B from the hepatopancreas of northern shrimp Pandalus borealis. Comparative Biochemistry and Physiology, 134B, 681–694.

Ashie, I. N. A., & Simpson, B. K. (1997). Proteolysis in food myosistem – a review. Journal of Food Biochemistry, 21, 91–123.

Boetius, A., & Felbeck, H. (1995). Digestive enzymes in marine invertebrates from hydrothermal vents and other reducing environments. Marine Biology, Berlin, Heidelberg, 122(1), 105–113.

Butler, A. M., Aiton, A. L., & Warner, A. H. (2001). Characterization of a novel heterodimeric cathepsin L-like protease and cDNA encoding the catalytic subunit of the protease in embryos of Artemia franciscana. Biochemistry and Cell Biology, 79, 43–56.

Dabrowski, K., & Glogowski, J. (1977). The role of exogenic proteolytic enzymes in digestion processes in fish. Hydrobiologia (Hagua), 54, 129–134.

Dendinger, J. E. (1987). Digestive proteases in the midgut gland of the Atlantic blue crab, Callinectes sapidus. Comparative Biochemistry and Physiology, 88B(2), 503–516.

Diaz-Tenorio, L. M., Garcia-Carreňo, F. L., & Navarrete del Toro, M. Á. (2006). Characterization and comparison of digestive proteinases of the Cortez swimming crab, Callinectes bellicosus, and the arched swimming crab, Callinectes arcuatus. Invertebrate Biology, 125(2), 125–135.

Dittrich, B. (1990). Temperature dependence of the activities of trypsin-like proteases in decapod crustaceans from different habitats. Naturwissenschaften, 77, 491–492.

Dittrich, B. (1992). Comparative studies on the thermal properties of a trypsin-like protease in two hermit crabs. Helgoländer Meeresuntersuchunger: Helgoländer Meeresunters, 46, 45–52.

Dixon, M., & Webb, E. C. (1964). Enzymes, 2rd ed. Longmans, Green & Co., London, New York.

Duan, Y. F., Liu, P., Li, J. T., Li, J., Gao, B. Q., & Chen, P. (2013). Cloning and expression analysis of cathepsin L cDNA of Exopalaemon carinicauda. Dongwuxue Yanjiu, 34(1), 39–46.

Egorova, V. V., Iezuitova, N. N., Timofeeva, N. M., Tulyaganova, E. K., Gurman, E. G., Shcherbakov, G. G., & Ugolev, A. M. (1974). Nekotoryye temperaturnye kharakteristiki i temperaturnye adaptatsii fermentov, obespechivayushchikh membrannoye pishchevareniye u poykilotermnykh i gomoyotermnykh zhivotnykh [Some temperature characteristics and temperature adaptations of enzymes that provide membrane digestion in poikilothermic and homeothermic animals]. Journal of Evolutionary Biochememistry and Physiology, 10(3), 223–231 (in Russian).

Fernández-Gimenez, A. V., García-Carreňo, F. L., Navarrete del Toro, M. A., & Fenucci, J. L. (2001). Digestive proteinases of red shrimp Pleoticus muelleri (Decapoda, Penaeoidea): Partial characterization and relationship with molting. Comparative Biochemistry and Physiology, 130B, 331–338.

Gerasimov, U. V. (Ed.). (2015). Ryba Rybinskogo vodokhranilishcha: Dinamika chislennosti naseleniya i ekologiya [Fish of the Rybinsk Reservoir: Population dynamics and ecology]. Filigran’, Yaroslavl (in Russian).

Gildberg, A. (1988). Aspartic proteinases in fishes and aquatic invertebrates. Comparative Biochemistry and Physiology, 91B, 425–435.

Glass, H. J., MacDonald, N. L., Moran, R. M., & Stark, J. R. (1989). Digestion of protein in different marine species. Comparative Biochemistry and Physiology, 94 B, 607–611.

Gerking, S. D. (1994). Feeding ecology of fish. Acadtmic Press, San-Diego.

Hemici, A., Benerbaiha, R. S., & Bendjeddou, D. (2017). Purification and biochemical characterization of a 22-kDa stable cysteine-like protease from the excretory-secretory product of the liver fluke Fasciola hepatica by using conventional techniques. Journal of Chromatography, B. Analytical Technologies in the Biomedical and Life Sciences Journal, 1068–1069, 268–276.

Hu, K.-J., & Leung, P.-C. (2007). Food digestion by cathepsin L and digestion-related rapid cell differentiation in shrimp hepatopancreas. Comparative Biochemistry and Physiology, 146B, 69–80.

Kim, H. R., Meyers, S. P., Pyeun, J. H., & Godber, J. S. (1994). Enzymatic properties of anionic trypsins from the hepatopancreas of crayfish, Procambarus clarkii. Comparative Biochemistry and Physiology, 107B, 197–203.

Kumar, S., Srivastava, A., & Chakrabarti, R. (2005). Study of digestive proteinases and proteinase inhibitors of Daphnia carinata. Aquaculture, 243, 367–372.

Kuz’mina, V. V. (1999). Vliyaniye temperatury na pishchevaritel'nyye gidrolazy vodnykh bespozvonochnykh [Influence of temperature on digestive hydrolases in water invertebrates]. Journal of Evolutionary Biochememistry and Physiology, 35(1), 15–19 (in Russiuan).

Kuz’mina, V. V. (2008). Classical and modern conceptions of fish digestion. In: Cyrino, J. E. P., Bureau, D. P., & Kapoor, B. G. (Eds.). Feeding and digestive functions in fishes. Ch. 4. Science Publications, Enfield (NF), Jersey, Plymouth. Pp. 85–154.

Kuz’mina, V. (2017). Digestion in fish. A new view. Lambert Academic Publishing, Balti.

Kuz’mina, V. V., Zolotareva, G. V., & Sheptitskiy, V. A. (2017). Proteolytic activity in some freshwater animals and associated microbiota in a wide pH range. Fish Physiology and Biochemistry, 43(2), 373–383.

Kuz’mina, V. V., Skvortsova, E. G., Shalygin, M. V., & Kovalenko, K. (2015). Role of peptidases of the enteral microbiota and preys in temperature adaptations of the digestive system in planktivorous and benthivorous fish. Fish Physiology and Biochemistry, 41(6), 1359–1368.

Lauff, M., & Hofer, R. (1984). Proteolytic enzymes in fish development and the importance of dietary enzymes. Aquaculture, 37(4), 335–346.

Le Boulay, C., Van Wormhoudt, A., & Sellos, D. (1996). Cloning and expression of cathepsin L-like proteinases in the hepatopancreas of the shrimp Penaeus vannamei during the intermolt cycle. Journal of Comparative Physiology, 166B(5), 310–318.

Li, X., Meng, X., Kong, J., Luo, K., Luan, S., Cao, B., Liu, N., Pang, J., & Shi, X. (2013). Molecular cloning and characterization of a cathepsin B gene from the Chinese shrimp Fenneropenaeus chinensis. Fish and Shellfish Immunology, 35(5), 1604–1612.

Munilla-Moran, R., Stark, J. R., & Babour, A. (1990). The role of exogenous enzymes in digestion in cultured turbot larvae (Scophthalmus maximus L.). Aquaculture, 88, 337–350.

Navarrete del Toro, M. A., García-Carreño, F. L., Díaz, L. M., Celis-Guerrero, L., & Saborowski, R. (2006). Aspartic proteinases in the digestive tract of marine decapod crustaceans. Journal of Experimental Zoology, 305A, 645–654.

Oh, E.-S., Kim, D.-S., Kim, J. H., & Kim, H.-R. (2000). Enzymatic properties of a protease from the hepatopancreas of shrimp, Penaeus oriantalis. Journal of Food Biochemistry, 24, 251–264.

Qiu, L., Jiang, S., Huang, J., Wang, W., Zhang, D., Wu, Q., & Yang, K. (2008). Molecular cloning and mRNA expression of cathepsin C gene in black tiger shrimp (Penaeus monodon). Comparative Biochemistry, 150A(3), 320–325.

Reid, R. G. B., & Rauchert, K. (1972). Protein digestion in members of the genus Macoma (Mollusca: Bivalvia). Comparative Biochemistry and Physiology, 41A, 887–895.

Reid, R. G. B., & Rauchert, K. (1976). Catheptic endopeptidases and protein digestion in the horse clam Tresus capax (Gould). Comparative Biochemistry and Physiology, 54, 467–472.

Sakai, J., Sakaguchi, Y., & Matsumoto, J. J. (1981). Acid proteinase activity of squid mantle muscle: Some properties. Comparative Biochemistry and Physiology, 70, 791–794.

Serviere-Zaragoza, E., Navarrete del Toro, M. A., & Garcia-Carreno, F. L. (1997). Protein hydrolyzing enzymes in the digestive systems of the adult Mexican blue abalone, Haliotis fulgens (Gastropoda). Aquaculture, 157, 323–332.

Skvortsova, E. G., Egorova, A. A., & Kuz’mina, V. V. (2016). Vliyaniye temperatury i kislotnosti sredy na aktivnost' peptidaz u lichinok khironomid – potentsial ob’yektov pitaniya ryb-bentofagov [Influence of the temperature and acidity of the medium on the activity of peptidases in larvae of chironomids – potential feeding objects of benthophagous]. Problems of Biology of Productive Animals, 4, 46–56 (in Russiuan).

Sriket, C., Benjakul, S., & Visessanguan, W. (2011). Characterisation of proteolytic enzymes from muscle and hepatopancreas of fresh water prawn (Macrobrachium rosenbergii). Journal of the Science of Food and Agriculture, 91(1), 52–59.

Stephens, A., Rojo, L., Araujo-Bernal, S., Garcia-Carreño, F., & Muhlia-Almazan, A. (2012). Cathepsin B from the white shrimp Litopenaeus vannamei: cDNA sequence analysis, tissues-specific expression and biological activity. Comparative Biochemistry and Physiology, 161B(1), 32–40.

Teschke, M., & Saborowski, R. (2005). Cysteine proteinases substitute for serine proteinases in the midgut glands of Crangon crangon and Crangon allmani (Decapoda: Caridea). Journal of Experimental Marine Biology and Ecology, 316, 213–229.

Van Wormhoudt, A., Sellos, D., Donval, A., Plaire-Goux, S., & Le Moullac, G. (1995). Chymotrypsin gene expression during the intermolt cycle in the shrimp Penaeus vannamei (Crustacea; Decapoda). Experientia, 51(2), 159–163.

Vega-Villasante, F., Nolasco, H., & Civera, R. (1995). The digestive enzymes of the Pacific brown shrimp Penaeus californiensis. II. Properties of protease activity in whole digestive tract. Comparative Biochemistry and Physiology, 112B, 123–129.

Wang, S., Shi, L. J., Liu, N., Chen, A. J., Zhao, X. F., & Wang, J. X. (2012). Involvement of Fenneropenaeus chinensis cathepsin C in antiviral immunity. Fish and Shellfish Immunology, 33(4), 821–828.

Warner, A. H., Pullumbi, E., Amons, R., & Liu, L. (2004). Characterization of a cathepsin L-associated protein in Artemia and its relationship to the FAS-I family of cell adhesion proteins. European Journal of Biochemistry, 271, 4014–4025.

Zhao, W., Chen, L., Zhang, F., Wu, P., Li, E., & Qin, J. (2013). Molecular characterization of cathepsin L cDNA and its expression during oogenesis and embryogenesis in the oriental river prawn Macrobrachium nipponense (Palaemonidae). Genetics and Molecular Research, 12(4), 5215–5225.


Most read articles by the same author(s)