Allocati N, Casalone E, Masulli M, Polekhina G, Rossjohn J, Parker M W, Di Ilio C
Dipartimento di Scienze Biomediche, Università 'G. D'Annunzio', Via dei Vestini 31, I-66013 Chieti, Italy.
Biochem J. 2000 Oct 15;351 Pt 2(Pt 2):341-6.
Glutathione S-transferases (GSTs) normally use hydroxy-group-containing residues in the N-terminal domain of the enzyme for stabilizing the activated form of the co-substrate, glutathione. However, previous mutagenesis studies have shown that this is not true for Beta class GSTs and thus the origin of the stabilization remains a mystery. The recently determined crystal structure of Proteus mirabilis GST B1-1 (PmGST B1-1) suggested that the stabilizing role might be fulfilled in Beta class GSTs by one or more residues in the C-terminal domain of the enzyme. To test this hypothesis we mutated His(106) and Lys(107) of PmGST B1-1 to investigate their possible role in the enzyme's catalytic activity. His(106) was mutated to Ala, Asn and Phe, and Lys(107) to Ala and Arg. The effects of the replacement on the activity, thermal stability and antibiotic-binding capacity of the enzyme were examined. The results are consistent with the involvement of His(106) and Lys(107) in interacting with glutathione at the active site but these residues do not contribute significantly to catalysis, folding or antibiotic binding.
谷胱甘肽S-转移酶（GSTs）通常利用酶N端结构域中含羟基的残基来稳定共底物谷胱甘肽的活化形式。然而，先前的诱变研究表明，β类GSTs并非如此，因此这种稳定作用的起源仍是个谜。奇异变形杆菌GST B1-1（PmGST B1-1）最近确定的晶体结构表明，β类GSTs的稳定作用可能由酶C端结构域中的一个或多个残基来实现。为了验证这一假设，我们将PmGST B1-1的His(106)和Lys(107)进行突变，以研究它们在酶催化活性中的可能作用。His(106)突变为Ala、Asn和Phe，Lys(107)突变为Ala和Arg。检测了这些替换对酶活性、热稳定性和抗生素结合能力的影响。结果表明His(106)和Lys(107)参与了在活性位点与谷胱甘肽的相互作用，但这些残基对催化、折叠或抗生素结合没有显著贡献。
