Ojennus Deanna Dahlke, Bratt Nicholas J, Jones Kent L, Juers Douglas H
Department of Chemistry, Whitworth University, 300 West Hawthorne Road, Spokane, WA 99251, USA.
Department of Mathematics and Computer Science, Whitworth University, 300 West Hawthorne Road, Spokane, WA 99251, USA.
Acta Crystallogr F Struct Biol Commun. 2019 Oct 1;75(Pt 10):625-633. doi: 10.1107/S2053230X19011774. Epub 2019 Sep 20.
Prolyl aminodipeptidase (PepX) is an enzyme that hydrolyzes peptide bonds from the N-terminus of substrates when the penultimate amino-acid residue is a proline. Prolyl peptidases are of particular interest owing to their ability to hydrolyze food allergens that contain a high percentage of proline residues. PepX from Lactobacillus helveticus was cloned and expressed in Escherichia coli as an N-terminally His-tagged recombinant construct and was crystallized by hanging-drop vapor diffusion in a phosphate buffer using PEG 3350 as a precipitant. The structure was determined at 2.0 Å resolution by molecular replacement using the structure of PepX from Lactococcus lactis (PDB entry 1lns) as the starting model. Notable differences between the L. helveticus PepX structure and PDB entry 1lns include a cysteine instead of a phenylalanine at the substrate-binding site in the position which confers exopeptidase activity and the presence of a calcium ion coordinated by a calcium-binding motif with the consensus sequence DX(DN)XDG.
脯氨酰氨基二肽酶（PepX）是一种酶，当倒数第二个氨基酸残基为脯氨酸时，它能从底物的N端水解肽键。脯氨酰肽酶因其能够水解含有高比例脯氨酸残基的食物过敏原而备受关注。来自瑞士乳杆菌的PepX被克隆并在大肠杆菌中作为N端带有His标签的重组构建体进行表达，并通过悬滴气相扩散法在以聚乙二醇3350作为沉淀剂的磷酸盐缓冲液中结晶。使用乳酸乳球菌PepX的结构（PDB编号1lns）作为起始模型，通过分子置换法在2.0 Å分辨率下确定了该结构。瑞士乳杆菌PepX结构与PDB编号1lns之间的显著差异包括：在赋予外肽酶活性的位置的底物结合位点处，一个半胱氨酸取代了苯丙氨酸，以及存在一个由具有一致序列DX(DN)XDG的钙结合基序配位的钙离子。
