Bacillus ComQ participates in the biosynthesis of a quorum-sensing signaling molecule (ComX pheromone) through catalyzing the prenylation at a Trp residue of the precursor peptide (pre-ComX) with geranyl diphosphate (C10 type) or farnesyl diphosphate (C15 type). We hypothesized that several residues specifically conserved among either type of ComQs are important for their substrate specificities. Using a simple bioassay, we revealed that Phe63, Asn186, and Gly190 in ComQRO-E-2 (C10 type) were nondisplaceable to Ser63, Gly186, and Val190, the corresponding residues in the C15-type ComQ, respectively. A three-dimensional model suggested that the 186th and 190th residues are involved in the pre-ComX binding. In vitro analysis showed that substitution of Phe63 with Ser in ComQRO-E-2 significantly reduced the geranylation activity but substantially enhanced the farnesylation activity, whereas substitution of Ser63 with Phe in ComQ168 (C15 type) reduced the farnesylation activity. Therefore, the 63rd residue was found to be significant for the prenyl-substrate preference.Abbreviations: GPP: geranyl diphosphate; FPP: farnesyl diphosphate; IPP: isopentenyl diphosphate; GGPP: geranylgeranyl diphosphate; FARM: first aspartate-rich motif; SARM: second aspartate-rich motif; β-Gal: β-galactosidase; TBABG: tryptose blood agar base supplemented with glucose; X-gal: 5-bromo-4-chloro-3-indolyl-β-D-galactoside.

中文翻译：

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通过使用简单的生物分析系统，鉴定ComQ的催化活性的关键残基，ComQ是一种用于群体感应的芽孢杆菌异戊二酸化酶。

芽孢杆菌ComQ通过催化在前体肽（pre-ComX）的Trp残基上以香叶基二磷酸（C10型）或法呢基二磷酸（C15型）催化异戊二烯参与到群体感应信号分子（ComX信息素）的生物合成中。我们假设两种类型的ComQ中特别保守的几个残基对其底物特异性很重要。使用简单的生物测定，我们揭示了ComQRO-E-2（C10型）中的Phe63，Asn186和Gly190分别不可替代Ser15，Gly186和Val190（C15型ComQ中的相应残基）。三维模型表明，第186和第190个残基参与了Pre-ComX的结合。体外分析表明，在ComQRO-E-2中用Ser取代Phe63显着降低了香叶基化活性，但大大增强了法呢基化活性，而在ComQ168（C15型）中用Phe取代Ser63则降低了法呢基化活性。因此，发现第63个残基对于异戊二烯基底物偏好是重要的。FPP：二磷酸法呢酯；IPP：二磷酸异戊烯基；GGPP：香叶基香叶基二磷酸；农场：第一个富含天冬氨酸的基序；SARM：第二种富含天冬氨酸的基序；β-Gal：β-半乳糖苷酶；TBABG：补充葡萄糖的类胰蛋白酶血琼脂基质；X-gal：5-溴-4-氯-3-吲哚基-β-D-半乳糖苷。发现第63个残基对于异戊二烯-底物偏好是重要的。FPP：二磷酸法呢酯；IPP：二磷酸异戊烯基；GGPP：香叶基香叶基二磷酸；农场：第一个富含天冬氨酸的基序；SARM：第二种富含天冬氨酸的基序；β-Gal：β-半乳糖苷酶；TBABG：补充葡萄糖的类胰蛋白酶血琼脂基质；X-gal：5-溴-4-氯-3-吲哚基-β-D-半乳糖苷。发现第63个残基对于异戊二烯-底物偏好是重要的。FPP：二磷酸法呢酯；IPP：二磷酸异戊烯基；GGPP：香叶基香叶基二磷酸；农场：第一个富含天冬氨酸的基序；SARM：第二种富含天冬氨酸的基序；β-Gal：β-半乳糖苷酶；TBABG：补充葡萄糖的类胰蛋白酶血琼脂基质；X-gal：5-溴-4-氯-3-吲哚基-β-D-半乳糖苷。