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Abstract The lepidopteran-active Cry1A.105 protein is a chimeric three-domain insecticidal toxin with distinct structural domains derived from the naturally occurring Cry1Ab, Cry1Ac and Cry1F proteins from the soil bacterium Bacillus thuringiensis (Bt). The X-ray crystal structure of the Cry1A.105 tryptic core at 3.0 Å resolution demonstrates its high structural similarity to the tryptic core of Cry1Ac. Bioinformatics analyses demonstrate that Cry1A.105 has no significant amino acid sequence similarity to known allergens or mammalian toxins, which is the same conclusion reached for its component domains. Like its intact donor proteins, Cry1A.105 was heat labile at temperatures ≥75 °C and degraded upon exposure to gastrointestinal proteases. No adverse effects were observed in mice when Cry1A.105 was dosed orally at 2451 mg/kg body weight. Therefore, the weight of evidence supports that Cry1A.105 is safe for human and animal consumption. These results support the conclusion that the safety of a chimeric protein for human or animal consumption can be evaluated in the context of the safety of its donor proteins. Highlights • Cry1A.105 is an engineered chimeric three-domain insecticidal protein. • Cry1A.105 displays the similar safety profiles to its donor proteins. • The domain exchange for creating Cry1A.105 preserves the Cry1 mode of action. • The weight of evidence supports food and feed safety of the soybean Cry1A.105 • HOSU of donor proteins is relevant to safety assessment of a chimeric protein. [ABSTRACT FROM AUTHOR]

Abstract: The phospholipase D (PLD) family of Arabidopsis thaliana has 12 identified members, including three highly homologous PLDγs. The enzymatic and molecular properties of PLDγ2 were characterized and compared with those of PLDγ1. Two variants of PLDγ2 cDNAs, designated PLDγ2a and PLDγ2b, were isolated, and they differ in the presence of a 96-nucleotide fragment at the beginning of the open reading frame. Catalytically active PLDγ2a was expressed in E. coli. PLDγ2a and γ1 both require phosphatidylinositol 4,5-bisphosphate (PIP2) and calcium for activity, but they differ in the effect of PIP2 and Triton X-100 on their activities. While Triton X-100 could greatly activate PLDγ1 activity and served only as a neutral diluent in the substrate vesicles, it totally abolished PLDγ2a activity and prohibited any stimulation effect from PIP2. PLDγ2a misses one of the basic, PIP2-interacting residues, which may weaken the binding of PIP2 to PLDγ2a. In addition, PLDγ2 and PLDγ1 displayed different patterns of expression in different tissues, and the transcript of PLDγ2a differs from that of PLDγ1 by having a longer 5′-UTR. These differences in biochemical and molecular properties suggest that the highly homologous PLDγs are subjected to unique regulations and might have distinguishable functions. [Copyright &y& Elsevier]