Crystallographic studies of Helicobacter pylori chaperone HspB and human serum transferrin : metalloprotein as a template for heavy metal ions and their relevance to bismuth antiulcer drug

Abstract

Iron is important for human health and serves as a co-factor in a variety of proteins and enzymes. Human serum transferrin (hTF) is an Fe(III) transporter in blood plasma which delivers metal to cells via a receptor-mediated endocytosis.

In the first part, crystal structures of FeNFeC-hTF and BiNFeC-hTF have been characterized. The N-lobes of the two structures adopt “partially opened” conformations between holo-hTF’s “closed” and apo-hTF’s “fully-opened” states. The N-lobe of BiNFeC-hTF opens wider than FeNFeC-hTF. Their metal-bound C-lobes are totally closed. Rigid-body movement and different inter-lobal hydrogen bonds for the “partially opened” conformations are observed. The binding affinities of four putative binding residues are in the order: Tyr188>Tyr95>Asp63~His249. In the N-lobe of BiNFeC-hTF, Tyr188, bicarbonate and a nitrilotriacetate (NTA) ion bind to Bi(III), whilst Tyr95 and Asp63 interact with NTA ligand. One (BiNFeC-hTF) or two (FeNFeC-hTF) glycan molecules are identified on the surface area of C-lobe. In the second part, biocoordination chemistry of selected metal ions was investigated using hTF as a template. The Al(III), Fe(III), Ga(III), Dy(III) and Yb(III)-bound hTF exhibit closed conformations in the C-lobe and “fully-opened” conformations in the N-lobe. In these structures, malonate serves as an anion in the C-lobe and provides two tunable ligation sites that lead to a less distorted octahedral coordination geometry. As a result, the large lanthanide ions (Dy(III) and Yb(III)) turn from their favored high coordination numbers (8~12) and fit into the protein’s hexadental pocket. Unexpectedly, in the presence of malonate ion and the excess amount of Dy(III) ion, the Ga(III) can be partially replaced by Dy(III), although Ga(III) has a much higher affinity than Dy(III) towards the protein. The chaperone system in Helicobacter pylorithat helps protein refold is assembled with HspB and HspA. In the third part, preliminary crystallographic work is reported for HspB and HspA. The chaperone HspB has been crystallized under various conditions and currently the diffraction resolution is 6.8Å. The co-chaperone HspA, which binds Bi(III) tightly, although its crystals diffract to 1.6Å, still needs improvement for data collection due to radiation damage.The crystal structure of HspB revealed that HspB presents as a single-ring heptamer, though it is a mixture of dimer, tetramer and a higher oligomer in solution. The interactions between HspB monomers in crystal structure are significantly weaker than that of GroEL (counterpart in Escherichia coli) monomers which may makes the HspB heptamer easier to dissociate in solution.