Purification and preparation of Rhodobacter sphaeroides reaction centers for photocurrent measurements and atomic force microscopy characterization

• Published in: STAR protocols 2022-03, Vol.3 (1), p.101044-101044, Article 101044
• Main Authors: Jun, Daniel, Zhang, Sylvester, Grzędowski, Adrian Jan, Mahey, Amita, Beatty, J. Thomas, Bizzotto, Dan
• Format: Article
• Language: English
• Subjects: Atomic Force Microscopy (AFM); Biophysics; Chemistry; Electrodes; Energy; Gold - chemistry; Material sciences; Microbiology; Microscopy; Microscopy, Atomic Force - methods; Photosynthetic Reaction Center Complex Proteins - chemistry; Physics; Protein Biochemistry; Protein expression and purification; Protocol; Rhodobacter sphaeroides - chemistry
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2021.101044

The formation of defined surfaces consisting of photosynthetic reaction centers (RCs) in biohybrid solar cells is challenging. Here, we start with the production of engineered RCs for oriented binding. RCs are deposited onto gold electrodes, and 6-mercapto-1-hexanol (MCH) is used to displace multilayers and non-specifically adsorbed RCs. The resulting electrode surfaces are analyzed for photocurrent generation using an intensity-modulated light and lock-in amplifier. Atomic force microscopy (AFM) is used to characterize the surface and the formation of RC structural assemblies. For complete details on the use and execution of this profile, please refer to Jun et al. (2021). [Display omitted] •Express and purify RC membrane proteins from Rhodobacter sphaeroides•Deposit RCs and MCH on gold electrodes for formation of structural assemblies•Measure RC photocurrents with an intensity-modulated LED and lock-in detection•Prepare single crystal gold electrodes for AFM of RCs and image processing The formation of defined surfaces consisting of photosynthetic reaction centers (RCs) in biohybrid solar cells is challenging. Here, we start with the production of engineered RCs for oriented binding. RCs are deposited onto gold electrodes, and 6-mercapto-1-hexanol (MCH) is used to displace multilayers and non-specifically adsorbed RCs. The resulting electrode surfaces are analyzed for photocurrent generation using an intensity-modulated light and lock-in amplifier. Atomic force microscopy (AFM) is used to characterize the surface and the formation of RC structural assemblies.