Thoden James B, Holden Hazel M
Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706, USA.
J Biol Chem. 2002 Jun 7;277(23):20854-61. doi: 10.1074/jbc.M201415200. Epub 2002 Mar 20.
Galactose mutarotase plays a key role in normal galactose metabolism by catalyzing the interconversion of beta-D-galactose and alpha-D-galactose. Here we describe the three-dimensional architecture of galactose mutarotase from Lactococcus lactis determined to 1.9-A resolution. Each subunit of the dimeric enzyme displays a distinctive beta-sandwich motif. This tertiary structural element was first identified in beta-galactosidase and subsequently observed in copper amine oxidase, hyaluronate lyase, chondroitinase, and maltose phosphorylase. Two cis-peptides are found in each subunit, namely Pro(67) and Lys(136). The active site is positioned in a rather open cleft, and the electron density corresponding to the bound galactose unequivocally demonstrates that both anomers of the substrate are present in the crystalline enzyme. Those residues responsible for anchoring the sugar to the protein include Arg(71), His(96), His(170), Asp(243), and Glu(304). Both His(96) and His(170) are strictly conserved among mutarotase amino acid sequences determined thus far. The imidazole nitrogens of these residues are located within hydrogen bonding distance to the C-5 oxygen of galactose. Strikingly, the carboxylate group of Glu(304) is situated at approximately 2.7 A from the 1'-hydroxyl group of galactose, thereby suggesting its possible role as a general acid/base group.
半乳糖变旋酶通过催化β-D-半乳糖和α-D-半乳糖的相互转化，在正常半乳糖代谢中发挥关键作用。在此，我们描述了乳酸乳球菌半乳糖变旋酶的三维结构，其分辨率达到1.9埃。二聚体酶的每个亚基都呈现出独特的β-折叠基序。这种三级结构元件最初在β-半乳糖苷酶中被鉴定出来，随后在铜胺氧化酶、透明质酸裂解酶、软骨素酶和麦芽糖磷酸化酶中也有观察到。每个亚基中发现了两个顺式肽段，即Pro(67)和Lys(136)。活性位点位于一个相当开放的裂隙中，与结合的半乳糖相对应的电子密度明确显示，底物的两种异头物都存在于晶体酶中。负责将糖锚定到蛋白质上的残基包括Arg(71)、His(96)、His(170)、Asp(243)和Glu(304)。His(96)和His(170)在迄今为止确定的变旋酶氨基酸序列中都是严格保守的。这些残基的咪唑氮原子位于与半乳糖C-5氧原子的氢键距离内。引人注目的是，Glu(304)的羧基位于距半乳糖1'-羟基约2.7埃处，因此表明其可能作为一般酸/碱基团发挥作用。
