Optimization of glycosidases production by Pseudoalteromonas issachenkonii KMM 3549 T Y.V. Alexeeva, 1,2 E. P. Ivanova, 2,3 I.Y. Bakunina, 2 T.N. Zvaygintseva 2 and V.V. Mikhailov 2 1 Far-Eastern State University, Vladivostok, Russia, 2 Pacific Institute of Bioorganic Chemistry of the Far-Eastern Branch of the Russian Academy of Sciences, Vladivostok, Russia, 3 Industrial Research Institute, Swinburne University of Technology, Vic., Australia 2002 ⁄ 47: received 7 February 2002, revised 21 May 2002 and accepted 26 June 2002 Y.V. ALEXEEVA, E. P. IVANOVA, I.Y. BAKUNINA, T.N. ZVAYGINTSEVA AND V.V. MIKHAILOV. 2002. Aims: The present work aimed to design an optimized medium to yield a higher production of glycosides by Pseudoalteromonas issachenkonii KMM 3549 T . Methods and Results: Higher levels of fucoidan hydrolase, alginase, laminaranase and b-N-acetylglucosaminidase production were obtained with peptone concentrations ranging from 2®5gl )1 to 10 g l )1 , while the presence of both yeast extract and glucose did not affect enzyme production. The activity of fucoidan hydrolase and laminaranase increased up to 4®83 lMh )1 mg )1 and 19®23 lMh )1 mg )1 protein, respectively, in growth media containing xylose (1®0gl )1 ), laminarin (0®5gl )1 ) or alginate (0®5gl )1 ), and production of b-N-acetylglucosaminidase substantially increased in the presence of fucoidan (0®5gl )1 ) or galactose (1 g l )1 ). All polysaccharides tested in concentrations of 0®5gl )1 fucoidan and 0®2gl )1 fucose induced production of alginase (up to 5®06 lMh )1 mg )1 protein). Conclusions: The production of glycosidases is not only stimulated by the presence of algal polysaccharides, but may also be stimulated by monosaccharides (e.g. xylose). Significance and Impact of the Study: The production of glycosidases by Pseudoalteromonas issachenkonii KMM 3549 T was significantly improved by using a simple nutrient medium containing peptone (2®5gl )1 ) and xylose (5®0gl )1 ) in 100% natural seawater. INTRODUCTION Microbial communities residing on the surfaces of macro- algae are characterized by wide-ranging physiological adap- tations and a high level of metabolic activity (Cho and Azam 1988; Warren 1996; Jaffray et al. 1997; Holmstro ¨m and Kjelleberg 1999). Recently we described the symbiotrophic associations of the marine aerobic gamma Proteobacteria, which are able to degrade the thallus of the brown algae Fucus evanescens (Ivanova et al. 2002a). A member of these associations assigned to the new species Pseudoalteromonas issachenkonii KMM 3549 T (Ivanova et al. 2002b) produced a number of glycosidases, namely fucoidan hydrolase, lamin- aranase, alginase, pustulanase, b-glucosidase, b-galactosi- dase, b-N-acetylglucosaminidase and b-xylosidase. This wide range of hydrolytic enzymes catalyse the hydrolysis of the main polysaccharides, e.g. alginic acids, cellulose, laminarans, and fucoidans of brown alga (Warren 1996). Laminarins (1,3-b-D-glucans) and its analogues were found in fungal cell walls, plants and lichens, while alginic acids and fucoidans were never detected in terrestrial sources. Fucoi- dans comprise an array of sulphated homo- and heteropoly- saccharides composed of a-(1(r)2)- and ⁄ or a-(1(r)3)-linked L-fucose residues (Patankar et al. 1993; Bilan et al. 2002) and also contain residues of galactose, mannose, xylose and glucuronic acid. The proportion of sulphated homo- and heteropolysaccharides as well as mono- and disaccharides varies greatly depending on both the species and geograph- ical origin of the brown algae (Nishino et al. 1991; Beress et al. 1993; Zvyagintseva et al. 1999). Because of such diversity, precise structures of fucoidans remain unclear. During the last decade studies of fucoidan structures received increasing attention as a result of their pharmacological Correspondence to: Elena P. Ivanova, Industrial Research Institute, Swineburne University of Technology, PO Box 218, Hawthorn, Vic 3122, Australia (e-mail: eivanova@swin.edu.au). ª 2002 The Society for Applied Microbiology Letters in Applied Microbiology 2002, 35, 343–346