Differentiation of Glycanases of Microbial Cellulolytic Systems Using Chromogenic and Fluorogenic Substrates

Abstract

Microbial cellulolytic systems are composed of several enzyme components attacking cellulose, cello-oligosaccharides and xylan. An understanding of the nature of various enzymes and their substrate specificities is important in developing processes that have industrial potential. Reliable enzyme differentiation is needed especially if genes for some enzymes are to be cloned. This need has been recently partially filled by introduction of sensitive methods for detection and identification of endo-1,4-β-glucanases and endo-1,4-β-xylanases employing the binding of Congo Red to carboxymethylcellulose and xylan [1–3], or employing the soluble covalently dyed hydroxyethylcellulose [4] and xylan [4,5]. Additional progress in the area of differentiation of cellulolytic enzymes has been achieved by the use of several fluorogenic substrates, 4-methylumbelliferyl β-glycosides of glucose, cellobiose [6] and lactose [6,7]. In this contribution we would like to report that a suitable combination of chromogenic and fluorogenic substrates for the detection of enzyme activities in separation gels (after electrophoresis or isoelectric focusing) or in solid growth selection media, permits a rapid and efficient differentiation of glycanases and some glycosidases according to their substrate specificities.