Article
Metal- and Metallocycle-Binding Sites Engineered into Polyvalent Virus-Like Scaffolds
Bioconjugate Chemistry
(2010)
Abstract
Metal-binding motifs appear on protein scaffolds throughout nature and are critical for a vast array of functions that span structure, electron transfer, and catalysis. In an effort to reproduce and exploit this activity in vitro, described herein are versatile bacteriophage Qβ particles bearing metal-binding motifs polyvalently. The three motifs, His6, His6-His6, and Cys-His6, were incorporated into the capsid via a coexpression methodology at ratios of 1.1:1, 1.1:1, and 2.3:1 for wild-type to modified coat protein. Size-exclusion chromatography yielded elution profiles identical to wild-type particles, while Ni-NTA affinity chromatography resulted in retention times that increase according to Qβ-His6 < Qβ-Cys-His6 < Qβ-His6-His6. In addition to interacting with metal-derivatized surfaces, Qβ-Cys-His6 and Qβ-His6-His6 bind heme as evidenced by the appearance of new absorbances at 416 and 418 nm, respectively, upon addition of hemin-Cl. The heme-bearing particles were also found to be electrochemically active as a surface-confined system. While both constructs yield similar E1/2 values anaerobically and with carbon monoxide present, and both display similar pH dependences, a standard rate constant k° could only be measured for Qβ-Cys-His6 (83 s−1), as electron transfer for Qβ-His6-His6 was too rapid to estimate. Experiments with rotated-disk electrodes yielded significant activity of the constructs toward dioxygen reduction. The versatility of the particles is further underscored by their multivalent nature, permitting simultaneously a range of activities for applications demanding polyfunctionality.
Disciplines
Publication Date
January 21, 2010
DOI
10.1021/bc900399e
Citation Information
Andrew K Udit, William Hollingsworth and Kang Choi. "Metal- and Metallocycle-Binding Sites Engineered into Polyvalent Virus-Like Scaffolds" Bioconjugate Chemistry Vol. 21 Iss. 2 (2010) p. 399 - 404 Available at: http://works.bepress.com/udit/36/