Hwang Peter M, Choy Wing-Yiu, Lo Eileen I, Chen Lu, Forman-Kay Julie D, Raetz Christian R H, Privé Gilbert G, Bishop Russell E, Kay Lewis E
Departments of Biochemistry, Medical Genetics and Microbiology, Laboratory Medicine and Pathobiology, and Chemistry, University of Toronto, Toronto, Ontario, Canada M5S 1A8.
Proc Natl Acad Sci U S A. 2002 Oct 15;99(21):13560-5. doi: 10.1073/pnas.212344499. Epub 2002 Sep 30.
The bacterial outer membrane enzyme PagP transfers a palmitate chain from a phospholipid to lipid A. In a number of pathogenic Gram-negative bacteria, PagP confers resistance to certain cationic antimicrobial peptides produced during the host innate immune response. The global fold of Escherichia coli PagP was determined in both dodecylphosphocholine and n-octyl-beta-d-glucoside detergent micelles using solution NMR spectroscopy. PagP consists of an eight-stranded anti-parallel beta-barrel preceded by an amphipathic alpha helix. The beta-barrel is well defined, whereas NMR relaxation measurements reveal considerable mobility in the loops connecting individual beta-strands. Three amino acid residues critical for enzymatic activity localize to extracellular loops near the membrane interface, positioning them optimally to interact with the polar headgroups of lipid A. Hence, the active site of PagP is situated on the outer surface of the outer membrane. Because the phospholipids that donate palmitate in the enzymatic reaction are normally found only in the inner leaflet of the outer membrane, PagP activity may depend on the aberrant migration of phospholipids into the outer leaflet. This finding is consistent with an emerging paradigm for outer membrane enzymes in providing an adaptive response toward disturbances in the outer membrane.
细菌外膜酶PagP将一条棕榈酸链从磷脂转移至脂多糖。在许多致病性革兰氏阴性菌中，PagP赋予对宿主固有免疫反应过程中产生的某些阳离子抗菌肽的抗性。利用溶液核磁共振波谱法，在十二烷基磷酸胆碱和正辛基-β-D-葡萄糖苷去污剂胶束中测定了大肠杆菌PagP的整体折叠结构。PagP由一个八链反平行β桶结构组成，前面有一个两亲性α螺旋。β桶结构明确，而核磁共振弛豫测量显示连接各个β链的环具有相当大的流动性。对酶活性至关重要的三个氨基酸残基定位于靠近膜界面的细胞外环，使其能够最佳地与脂多糖的极性头部相互作用。因此，PagP的活性位点位于外膜的外表面。由于在酶促反应中提供棕榈酸的磷脂通常仅在外膜的内膜层中发现，PagP的活性可能取决于磷脂异常迁移到外膜层。这一发现与外膜酶对外膜干扰提供适应性反应的新范式一致。
