Expression of Gene for β-glucanase from Paenibacillus jamilae Bg1 in Pichia pastoris and Characteristics of Recombinant Enzyme

Abstract

The isolation, heterologous expression and characterization of a new thermostable β-glucanase from Paenibacillus jamilae is described. The bgl26 gene from the P. jamilae Bg1 VKPM B-13093 strain consisting of 714 nucleotides encodes endo-1,3-1,4-β-glucanase (EC 3.2.1.73) containing 213 amino acids and 24 residues of the putative signal peptide in N-end area. The nucleotide sequence of the bgl26 gene and the amino acid sequence of the mature Bgl26 protein have the greatest homology with the sequence of the Paenibacillus macerans endo-l,3-l,4-β-glucanase (82 and 88%, respectively). A fragment of the gene encoding the mature protein was expressed in Pichia pastoris. Purified recombinant enzyme Bgl26 was active towards barley β-glucan. The optimal pH for the enzyme to work was 7,0, and the optimum temperature range was 40-45 °C. The specific activity of β-glucanase was at the level of 6650 U/mg of protein, Km and Vmax were equal to 6.4 ± 0.3 mg/mL and 9450.1 ± 471.2 umol/(min-mg), respectively. The recombinant protein Bgl26 was characterized by high pH and thermal stability, as well as resistance to digestive enzymes. It is also shown that Co2+ ions have a positive effect on the activity of the enzyme. β-glucanase, β-glucan, Paenibacillus jamilae, Pichia pastoris The work was financially supported by the Ministry of Science and Higher Education of the Russian Federation (Unique Project Identifier RFMEFI60717X0179) and was carried out using the Multipurpose Scientific Installation of National Bio-Resource Center «All-Russian Collection of Industrial Microorganisms», NRC «Kurchatov Institute» - GOSNIIGENETIKA.