Identification of endophytic actinobacteria from Jerusalem artichoke and examination of inulinase gene and enzyme properties

Abstract

Endophytic actinobacteria isolated from Helianthus tuberosus L. (Jerusalem artichoke) demonstrate great potency in the production of biological function and compounds, including exo-inulinase activity. Exo-inulinase is of interest to the beverage industry, as this enzyme hydrolyzes inulin stored within the Jerusalem artichoke tuber into fructose syrup in a single step. This study aimed to identify and investigate the exo-inulinase-producing endophytic actinobacteria from four varieties of Jerusalem artichoke (HEL65, JA89, CNS2867 and JA102×JA89) in root, leaf and stem samples. In total, 78 isolates of endophytic actinobacteria were classified into 24 different groups based on morphology and then identified based on the 16S rRNA gene into 5 genera: Streptomyces (64.1%), Nocardia (28.2%), Actinosynnema (3.9%), Kribbella (2.6%) and Kineococcus (1.3%). The results of inulinase screening using polymerase chain reaction with degenerated primers showed the specific band at 400 bp of Streptomyces gancidicus EAH-R3 which conserved the protein of the exo-inulinase gene and had inulinase activity of 125 U/mL. This inulinase was confirmed to be an exo-inulinase by checking the end product of inulin hydrolysis using thin layer chromatography. The crude enzyme of S. gancidicus EAH-R3 had an optimum temperature and pH of 60οC and 6.0, respectively. Moreover, the crude inulinase was thermostable at 80οC for 2 hr with a relative activity of 94%. S. gancidicus EAH-R3, a new isolate of endophytic actinobacteria isolated from the plant with high inulin content, is a candidate for fructose production using inulin hydrolysis.