Members of the metallo-β-lactamase (MBL) superfamily of enzymes harbor a highly conserved αββα MBL-fold domain and were first described as inactivators of common β-lactam antibiotics. In humans, these enzymes have been shown to exhibit diverse functions, including hydrolase activity towards amides, esters, and thioesters. An uncharacterized member of the human MBL family, MBLAC2, was detected in multiple palmitoylproteomes, identified as a zDHHC20 S-acyltransferase interactor, and annotated as a potential thioesterase. In this study, we confirmed that MBLAC2 is palmitoylated and identified the likely S-palmitoylation site as Cys254. S-palmitoylation of MBLAC2 is increased in cells when expressed with zDHHC20 and MBLAC2 is a substrate for purified zDHHC20 in vitro. To determine its biochemical function, we tested the ability of MBLAC2 to hydrolyze a variety of small molecules and acylprotein substrates. MBLAC2 has acyl-CoA thioesterase activity with kinetic parameters and acyl-CoA selectivity comparable to acyl-CoA thioesterase 1 (ACOT1). Two predicted zinc-binding residues, Asp87 and His88 are required for MBLAC2 hydrolase activity. Consistent with a role in fatty acid metabolism in cells, MBLAC2 was cross-linked to a photoactivatable fatty acid in a manner that was independent of its S-fatty acylation at Cys254. Our study adds to previous investigations demonstrating the versatility of the MBL-fold domain in supporting a variety of enzymatic reactions.

中文翻译：

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含金属-β-内酰胺酶结构域的蛋白 2 (MBLAC2) 被 S-棕榈酰化，并表现出酰基辅酶 A 水解酶活性


金属-β-内酰胺酶 (MBL) 酶超家族的成员具有高度保守的 αββα MBL 折叠结构域，最初被描述为常见 β-内酰胺抗生素的灭活剂。在人类中，这些酶已被证明表现出多种功能，包括对酰胺、酯和硫酯的水解酶活性。在多个棕榈酰蛋白质组中检测到人类 MBL 家族的一个未表征的成员 MBLAC2，被鉴定为 zDHHC20 S-酰基转移酶相互作用蛋白，并注释为潜在的硫酯酶。在这项研究中，我们确认 MBLAC2 被棕榈酰化，并确定了可能的 S-棕榈酰化位点为 Cys254。当与 zDHHC20 一起表达时，MBLAC2 的 S-棕榈酰化在细胞中增加，并且 MBLAC2 是体外纯化的 zDHHC20 的底物。为了确定其生化功能，我们测试了 MBLAC2 水解各种小分子和酰基蛋白底物的能力。 MBLAC2 具有酰基辅酶 A 硫酯酶活性，其动力学参数和酰基辅酶 A 选择性与酰基辅酶 A 硫酯酶 1 (ACOT1) 相当。 MBLAC2 水解酶活性需要两个预测的锌结合残基 Asp87 和 His88。与细胞中脂肪酸代谢的作用一致，MBLAC2 以独立于 Cys254 处 S-脂肪酰化的方式与光活化脂肪酸交联。我们的研究补充了之前的研究，证明了 MBL 折叠结构域在支持各种酶促反应方面的多功能性。