A family 11 xylanase from Penicillium funiculosum is strongly inhibited by three wheat xylanase inhibitors

Steady-state kinetic approaches were used to investigate the binding of a novel Penicillium funiculosum xylanase, XYNC, with three known xylanase inhibitor proteins from wheat (Triticum aestivum). The xylanase gene (xynC) was cloned from a P funiculosum genomic library and the deduced amino acid sequence of XYNC exhibited high sequence similarity with fungal family 11 xylanases. xynC was overexpressed in P. funiculosum and the product (XYNC: M(r) = 23.6 kDa; pI = 3.7) purified and shown to efficiently degrade birchwood xylan [K(m) = 0.47% w/v, V(max) = 2540 mumol xylose min(-1) (mg protein)(-1) at pH 5.5 and 30 degreesC] and soluble wheat arabinoxylans [K(m) = 1.45% w/v, V(max) = 7190 mumol xylose min(-1) mg protein)(-1) at pH 5.5 and 30 degreesC]. The xylanase activity of XYNC was inhibited strongly by three xylanase inhibitor proteins from wheat; XIP-I, TAXI I and TAXI II. The inhibition for each was competitive, with very tight binding (K(i) = 3.4, 16 and 17 nM, respectively) equivalent to free energy changes (DeltaGdegrees) of - 49, - 45 and - 45 kJ mol(-1). This is the first report describing a xylanase that is inhibited by all three wheat xylanase inhibitor proteins described to date. (C) 2002 Elsevier Science B.V. All rights reserved.