Structural and biochemical properties of a novel pullulanase of Paenibacillus lautus DSM 3035

Abstract

The pullulanase gene (pulPL), encoding a novel type I pullulanase (PulPL), was obtained from a Paenibacillus lautus DSM3035 isolate. The gene has an open reading frame of 2355 bp, After optimizing induction conditions in Escherichia coli, we overexpressed recombinant PulPL, purified this enzyme, and assayed its function . The level of functional PulPL‐like protein reached its maximum (about 0.28 mg/mL, 15% of total protein) after induction for 16 h at 20°C. Under these optimized harvesting conditions, PulPL activity was 11.1 U/mL. The purified recombinant enzyme with an apparent molecular mass of about 87.9 kDa was able to specifically attack the α‐1,6 linkages in pullulan to generate maltotriose as the major product. The purified PulPL exhibited optimal activity at pH 7.0 and 40°C. The PulPL hydrolyzed pullulan, amylopectin, starch, and glycogen, but not amylose. Substrate specificity observations and reaction products identifications indicate that the purified pullulanase from P. lautus DSM3035 is a type I pullulanase. The present report, to our knowledge, the first to identify type I pullulanase in P.lautus, and detail the enzymatic properties of this enzyme after heterologous expression.