6DHZ

Crystallographic octamer of a metal-free RIDC1 variant bearing two disulfide bonded cysteines

Summary for 6DHZ

• Entry DOI: 10.2210/pdb6dhz/pdb
• Descriptor: Soluble cytochrome b562, HEME C, CALCIUM ION, ... (4 entities in total)
• Functional Keywords: 4-helix bundle, engineered protein, metalloprotein, electron transport
• Biological source: Escherichia coli
• Total number of polymer chains: 4
• Total formula weight: 49687.31

Authors

Tezcan, F.A.,Churchfield, L.A. (deposition date: 2018-05-21, release date: 2018-07-25, Last modification date: 2023-10-11)

Primary citation

Churchfield, L.A.,Alberstein, R.G.,Williamson, L.M.,Tezcan, F.A.
Determining the Structural and Energetic Basis of Allostery in a De Novo Designed Metalloprotein Assembly.
J. Am. Chem. Soc., 140:10043-10053, 2018

PubMed Abstract: Despite significant progress in protein design, the construction of protein assemblies that display complex functions (e.g., catalysis or allostery) remains a significant challenge. We recently reported the de novo construction of an allosteric supramolecular protein assembly (Zn-R1) in which the dissociation and binding of Zn ions were coupled over a distance of 15 Å to the selective hydrolytic breakage and formation of a single disulfide bond. Zn-R1 was constructed by Zn-templated assembly of a monomeric protein (R1, a derivative of cytochrome cb) into a tetramer, followed by progressive incorporation of noncovalent and disulfide bonding interactions into the protein-protein interfaces to create a strained quaternary architecture. The interfacial strain thus built allowed mechanical coupling between the binding/dissociation of Zn and formation/hydrolysis of a single disulfide bond (C38-C38) out of a possible six. While the earlier study provided structural evidence for the two end-states of allosteric coupling, the energetic basis for allosteric coupling and the minimal structural requirements for building this allosteric system were not understood. Toward this end, we have characterized the structures and Zn-binding properties of two related protein constructs (R1 and R1) which also possess C38-C38 disulfide bonds. In addition, we have carried out extensive molecular dynamics simulations of R1 to understand the energetic basis for the selective cleavage of the C38-C38 disulfide bond upon Zn dissociation. Our analyses reveal that the local interfacial environment around the C38-C38 bond is key to its selective cleavage, but this cleavage is only possible within the context of a stable quaternary architecture which enables structural coupling between Zn coordination and the protein-protein interfaces.

PubMed: 29996654
DOI: 10.1021/jacs.8b05812

Experimental method

X-RAY DIFFRACTION (2.8 Å)