title: Serpin polymerization is prevented by a hydrogen bond network that is centered on his-334 and stabilized by glycerol.
abstract: Polymerization of serpins commonly results from mutations in the shutter region underlying the bifurcation of strands 3 and 5 of the A-sheet, with entry beyond this point being barred by a H-bond network centered on His-334. Exposure of this histidine in antithrombin, which has a partially opened sheet, allows polymerization and peptide insertion to occur at pH 6 or less when His-334 will be predictably protonated with disruption of the H-bond network. Similarly, thermal stability of antithrombin is pH-dependent with a single unfolding transition at pH 6, but there is no such transition when His-334 is buried by a fully closed A-sheet in heparin-complexed antithrombin or in alpha(1)-antitrypsin. Replacement of His-334 in alpha(1)-antitrypsin by a serine or alanine at pH 7.4 results in the same polymerization and loop-peptide acceptance observed with antithrombin at low pH. The critical role of His-334 and the re-formation of its H-bond network by the conserved P8 threonine, on the full insertion of strand 4, are relevant for the design of therapeutic blocking agents. This is highlighted here by the crystallographic demonstration that glycerol, which at high concentrations blocks polymerization, can replace the P8 threonine and re-form the disrupted H-bond network with His-334.
authors.affiliation: Department of Haematology, University of Cambridge, Cambridge Institute of Medical Research, Hills Road, Cambridge CB2 2XY, United Kingdom. awz20@cus.cam.ac.uk
authors.forename: Aiwu
authors.initials: A
authors.lastname: Zhou
authors.forename: Penelope E
authors.initials: PE
authors.lastname: Stein
authors.forename: James A
authors.initials: JA
authors.lastname: Huntington
authors.forename: Robin W
authors.initials: RW
authors.lastname: Carrell
mesh_terms: D000991:Antithrombins / Q000737:chemistry; D018360:Crystallography, X-Ray; D005990:Glycerol / Q000494:pharmacology*; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D006860:Hydrogen Bonding / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D011108:Polymers; D011487:Protein Conformation / Q000187:drug effects; D011489:Protein Denaturation; D015843:Serpins / Q000737:chemistry* / Q000187:drug effects; D013696:Temperature; D000515:alpha 1-Antitrypsin / Q000737:chemistry
chemical_list: D000991:Antithrombins; D011108:Polymers; D015843:Serpins; D000515:alpha 1-Antitrypsin; D006639:Histidine; D005990:Glycerol