Yodsoi KanintronkulIssara SramalaGerd KatzenmeierSakol PanyimChanan AngsuthanasombatMahidol University2018-07-242018-07-242003-05-01Applied Biochemistry and Biotechnology - Part B Molecular Biotechnology. Vol.24, No.1 (2003), 11-19107360852-s2.0-0037952941https://repository.li.mahidol.ac.th/handle/20.500.14594/20730The widely accepted model for toxicity mechanisms of the Bacillus thuringiensis Cry δ-endotoxins suggests that helices α4 and α5 form a helix-loop-helix hairpin structure to initiate membrane insertion and pore formation. In this report, alanine substitutions of two polar amino acids (Asn-166 and Tyr-170) and one charged residue (Glu-171) within the α4-α5 loop of the 130-kDa Cry4B mosquito-larvicidal protein were initially made via polymerase chain reaction-based directed mutagenesis. As with the wild-type toxin, all of the mutant proteins were highly expressed in Escherichia coli as inclusion bodies upon isopropyl-β-D-thiogalactopyranoside induction. When E. coli cells expressing each mutant toxin were assayed against Aedes aegypti mosquito larvae, the activity was almost completely abolished for N166A and Y170A mutations, whereas E171A showed only a small reduction in toxicity. Further analysis of these two critical residues by induction of specific mutations revealed that polarity at position 166 and highly conserved aromaticity at position 170 within the α4-α5 loop play a crucial rote in the larvicidal activity of the Cry4B toxin.Mahidol UniversityBiochemistry, Genetics and Molecular BiologyChemical EngineeringImmunology and MicrobiologyArticleSCOPUS10.1385/MB:24:1:11