title: Phase changes in T(3)R(3)(f) human insulin: temperature or pressure induced?
abstract: The structure of T(3)R(3) hexameric human insulin has been determined at 100 K from two different crystals at 1.2 and 1.3 A resolution and refined to residuals of 0.169 and 0.176, respectively. Owing to a phase change, the c axis is double its room-temperature value and the asymmetric unit contains two independent TR(f) insulin dimers. Compared with the orientation in the room-temperature structure, one dimer undergoes a rotation about the c axis of -5 degrees, while the second is rotated +4 degrees. A superposition of the backbone atoms of the two independent dimers shows that the C(alpha) atoms of five residues within the R(f)-state monomers are displaced by more than 1.0 A; smaller displacements are observed for the T-state monomers. Four zinc ions lie on the crystallographic threefold axis and each forms bonds to three symmetry-related HisB10 N(varepsilon2) atoms from the T- and R(f)-state trimers. While three of the zinc ions are tetrahedrally coordinated with a chloride ion completing the coordination sphere, mixed tetrahedral/octahedral coordination is observed for one of the T-state zinc ions. The three symmetry-related "phenolic binding sites" in one hexamer contain water molecules and a glycerol molecule, but the same sites in the second hexamer are occupied by a zinc ion coordinated to an alternate conformation of HisB10, a symmetry-related HisB5 and two chloride ions. Two additional and partially occupied zinc ion sites are observed at the interface between the two independent dimers. One zinc ion is coordinated by a T-state HisB5 of one dimer, an R-state HisB5 of the second dimer and two water molecules; the second zinc ion is coordinated by an alternate side-chain conformation of the T-state HisB5 and three water molecules. The carboxyl group of one GluB13 side chain, which exists in two discrete conformations, appears to be protonated, because short contacts exist to a second carboxyl group or to a carbonyl O atom.
authors.affiliation: Hauptman-Woodward Medical Research Institute, 73 High Street, Buffalo, NY 14203, USA. smith@hwi.buffalo.edu
authors.forename: G D
authors.initials: GD
authors.lastname: Smith
authors.forename: W A
authors.initials: WA
authors.lastname: Pangborn
authors.forename: R H
authors.initials: RH
authors.lastname: Blessing
mesh_terms: D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D006801:Humans; D007328:Insulin / Q000737:chemistry*; D008958:Models, Molecular; D011108:Polymers / Q000737:chemistry; D011487:Protein Conformation; D015032:Zinc / Q000737:chemistry
chemical_list: D007328:Insulin; D011108:Polymers; D015032:Zinc