Cloning and expression of chitinase A, B, and C (chiA, chiB, chiC) genes from Serratia marcescens originating from Helicoverpa armigera and determining their activities

Authors : Mehtap DANIŞMAZOĞLU, İsmail DEMİR, Kazım SEZEN, Hacer MURATOĞLU, Remziye NALÇACIOĞLU

Pages : 78-87

Doi:10.3906/biy-1404-31

View : 10 | Download : 7

Publication Date : 2015-12-01

Article Type : Research Paper

Abstract :Three genes encoding chitinase A insert ignore into journalissuearticles values(chiA);, B insert ignore into journalissuearticles values(chiB);, and C insert ignore into journalissuearticles values(chiC); were amplified from a bacterium that was isolated from a naturally dead Helicoverpa armigera larva and identified as Serratia marcescens based on 16S rRNA gene sequence analysis insert ignore into journalissuearticles values(KF823633 accession number);. The open reading frames insert ignore into journalissuearticles values(ORFs); were identified as 1692, 1500, and 1443 base pairs for chiA, chiB, and chiC genes, respectively. These sequences were submitted to the GenBank with accession numbers KF823630 insert ignore into journalissuearticles values(chiA);, KF823631 insert ignore into journalissuearticles values(chiB);, and KF823632 insert ignore into journalissuearticles values(chiC);. Comparison of the deduced amino acid sequences with those of other bacterial chitinases revealed that the 3 chitinases contain the catalytic domain. Furthermore, all 3 chitinases showed 99% similarity to the S. marcescens WW4 strain at the amino acid level. The chitinases were overexpressed in Escherichia coli. Expressed proteins were purified and their activities were tested using colloidal chitin as substrate. Reasonable pH and temperature ranges were also determined as 7-11 and 33-37 °C, respectively. Insecticidal activities of these proteins were tested on the larvae of Malacosoma neustria and H. armigera. Test results showed that 1000 U/mL ChiA, ChiB, and ChiC have 47%, 50%, and 66% insecticidal activities on M. neustria, and 80%, 45%, and 50% insecticidal activities on H. armigera larvae within 10 days, respectively.
Keywords : Serratia marcescens, chiA, chiB, chiC, insecticidal activity