Formation and carbon monoxide‐dependent dissociation of Allochromatium vinosum cytochrome c′ oligomers using domain‐swapped dimers

Abstract

The number of artificial protein supramolecules has been increasing; however, control of protein oligomer formation remains challenging. Cytochrome c' from Allochromatium vinosum (AVCP) is a homodimeric protein in its native form, where its protomer exhibits a four-helix bundle structure containing a covalently bound five-coordinate heme as a gas binding site. AVCP exhibits a unique reversible dimer-monomer transition according to the absence and presence of CO. Herein, domain-swapped dimeric AVCP was constructed and utilized to form a tetramer and high-order oligomers. The X-ray crystal structure of oxidized tetrameric AVCP consisted of two monomer subunits and one domain-swapped dimer subunit, which exchanged the region containing helices αA and αB between protomers. The active site structures of the domain-swapped dimer subunit and monomer subunits in the tetramer were similar to those of the monomer subunits in the native dimer. The subunit-subunit interactions at the interfaces of the domain-swapped dimer and monomer subunits in the tetramer were also similar to the subunit-subunit interaction in the native dimer. Reduced tetrameric AVCP dissociated to a domain-swapped dimer and two monomers upon CO binding. Without monomers, the domain-swapped dimers formed tetramers, hexamers, and higher-order oligomers in the absence of CO, whereas the oligomers dissociated to domain-swapped dimers in the presence of CO, demonstrating that the domain-swapped dimer maintains the CO-induced subunit dissociation behavior of native ACVP. These results suggest that protein oligomer formation may be controlled by utilizing domain swapping for a dimer-monomer transition protein.

Conferment of CO-Controlled Dimer–Monomer Transition Property to Thermostable Cytochrome c′ by Mutation in the Subunit–Subunit Interface

“…The elution peak at ~11 mL of oxidized and reduced wild-type PHCP did not shift under CO-saturated reduced conditions, while the elution peak of oxidized AVCP at ~11 mL shifted to 12-13 mL under CO-saturated reduced conditions. 45 These results indicate that the wild-type PHCP dimer does not dissociate to monomers under CO-saturated reduced conditions that AVCP dissociates.…”

“…Recently, AVCP oligomers, which reversibly dissociate to dimers upon CO binding/dissociation, have been constructed by utilizing domain-swapped dimers. 45 In addition to a high CO affinity and a dimer-monomer transition property upon CO binding/dissociation for the T18F/F71D PHCP variant (Table 1 and Figure 3), its stability was higher than that of AVCP (Tm: T18F/F71D PHCP, 74.0 °C; AVCP, 52.4 °C) (Figure 4). These properties may be advantageous for construction of gas sensors and gas-controllable molecules.…”

New Aspects of Cytochrome c: 3D Domain Swapping, Membrane Interaction, Peroxidase Activity, and Met80 Sulfoxide Modification

“…54,55 Additionally, cyt cb 562 (a cyt b 562 mutant protein with a heme immobilized on the 4-helix bundle protein moiety by introducing two cysteine residues to the polypeptide), Shewanella violacea (SV) cyt c 5 , and Aquifex aeolicus cyt c¤ also domain swap and the protein structures of their 3D-DS dimers are similar to the corresponding structures of the monomers. 62,63 In these proteins, the hinge loop is located relatively far from the heme, resulting in the property of the dimer being similar to that of the corresponding monomer. The heme coordination structure and property of a heme protein may change by 3D-DS when the hinge loop is located close to the heme active site, but not when the hinge loop is located far from it.…”

Oligomerization of cytochrome c, myoglobin, and related heme proteins by 3D domain swapping

“…The sixth coordination site of AV cyt c′ is occupied with the side chain of Tyr16, which has been suggested to be a trigger for the dimer-to-monomer transition upon CO binding to the heme [89]. AV cyt c′ formed a domain-swapped dimer, and the dimer was utilized to form a tetramer, comprising one domain-swapped dimer subunit and two monomer subunits (PDB ID: 5GYR; Figure 8A) [90]. The heme environments of the monomer and domain-swapped dimer subunits of the tetramer were similar to those of the native dimer (Figure 8B).…”