The effect of sunlight on the protein crystals from Bacillus thuringiensis var. kurstaki HD1 and NRD12: A Raman spectroscopic study

Abstract The Raman spectrum of the endotoxin crystals from Bacillus thuringiensis var. kurstaki NRD12 is found to resemble closely that for crystals of the strain kurstaki HD1 (P. R. Carey et al., 1986). Since the Raman data provide detailed molecular information on the crystal proteins, the protoxins of HD1 and NRD12 must be in similar conformations in their respective crystals. Estimation of the protoxin secondary structure in NRD12 crystals, using the amide III and amide I′ Raman spectral features, gave 57% unordered, 25% α-helix, and 18% β-structure. This is very similar to the result for HD1, as is the finding that the aromatic amino acid side chains of the NRD12 protoxin in the crystal appear to have widespread accessibility to water molecules. The effect of sunlight irradiation on dried NRD12 or HD1 crystals was studied by recording the Raman spectrum of the crystals before and after irradiation under a solar simulator whose spectral output is essentially identical to that of sunlight at the earth's surface. The only Raman spectral change detected following 40-hr irradiation was a 35% reduction of the intensity of features near 760 and 1555 cm −1 due to tryptophan sidechains, indicating that irradiation brings about widespread destruction of indole residues. Control experiments using α-chymotrypsin, which has prominent tryptophan features in its Raman spectrum, show that the destruction of tryptophan is not brought about by the direct photolysis of the indole residues by the sunlight photons. Since silkworm bioassays on crystals of HD1 irradiated for 40 hr under the solar simulator or by natural sunlight showed that the crystals had become atoxic, the present results highlight the role of tryptophan destruction in sunlight-mediated crystal inactivation.