title: Identification and characterization of the Trichoderma harzianum gene encoding alpha-1,3-glucanase involved in streptococcal mutan degradation.
abstract: alpha-1,3-Glucanases (mutanases) are currently of great interest due to their potential use in the field of dental care. These enzymes have been reported in several bacteria, yeasts and fungi, but up to now, characterization of this family of proteins has been relatively poor. In this study, we identify and characterize a mutanase gene from Trichoderma harzianum CCM F-340. Sequence analysis, on the nucleotide and amino acid levels reveals that this alpha-1,3-glucanase is highly homologous to alpha-1,3-glucanases from T harzianum isolate CBS 243.71 and T asperellum CECT 20539. T. harzianum CCM F-340 mutanase is a 634-aa residue protein with a calculated molecular mass of 67.65 kDa, composed of two distinct, highly conserved domains (a long N-terminal catalytic domain and a short C-terminal polysaccharide-binding domain) separated by a less conserved Pro-Ser-Thr-rich linker region. The mutanase gene was expressed in an E. coli BL21 (DE3) host, under the transcriptional control of T7 promoter. The purified enzyme migrated as a band of about 68 kDa after SDS-polyacrylamide gel electrophoresis, which coincided with the predicted size based on the amino acid sequence. Our data indicate that this enzyme is highly conserved in Trichoderma and can be produced in active form in such heterologous expression system.
authors.affiliation: Department of Industrial Microbiology, Institute of Microbiology and Biotechnology, Maria Curie-Skłdodowska University, Lublin, Poland. adrianw2@wp.pl
authors.forename: Adrian
authors.initials: A
authors.lastname: Wiater
authors.forename: Monika
authors.initials: M
authors.lastname: Janczarek
authors.forename: Małgorzata
authors.initials: M
authors.lastname: Pleszczyńska
authors.forename: Janusz
authors.initials: J
authors.lastname: Szczodrak
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004789:Enzyme Activation; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010802:Phylogeny; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013291:Streptococcus / Q000378:metabolism*; D014242:Trichoderma / Q000145:classification / Q000201:enzymology / Q000235:genetics*
chemical_list: D011994:Recombinant Proteins; D006026:Glycoside Hydrolases; C021780:exo-1,3-alpha-glucanase