Energy transfer between fusion biliproteins co-expressed with phycobiliprotein in Escherichia coli.
In cyanobacteria, phycobiliproteins (PBS) show excellent energy transfer among the chromophores absorbing over most of the visible. The energy transfers are used to study phycobilisome assembly and bioimaging. Using All4261GAF2(C81L) as energy donor, ApcE(1-240/Δ87-130) as energy acceptor, we co-expressed fusion protein ApcE(1-240/Δ87-130)::All4261GAF2(C81L) with phycobiliprotein in Escherichia Coli and studied the energy transfer between two protein domains. With N-terminal His6 tag, ApcE(1-240/Δ87-130)::All4261GAF2(C81L) cannot be purified by nickel-affinity column. We added six histidines in the C-terminal of ApcE(1-240/Δ87-130)::All4261GAF2(C81L) and co-expressed it with phycobiliprotein. ApcE(1-240/Δ87-130)::PCB-All4261GAF2(C81L)His6 was purified successfully and only singly chromophorylated at All4261GAF2(C81L)His6 domain. The singly chromophorylate ApcE(1-240/Δ87-130)::PCB-All4261GAF2(C81L)His6 was incubated with fresh PCB and the doubly chromophorylated PCB-ApcE(1-240/Δ87-130)::PCB-All4261GAF2(C81L)His6 was obtained. The double chromophored fusion protein absorbed light in the range of 615-660 nm, and fluoresced only at 668 nm. Photochemistry analysis showed that excitation energy transfer from the short-wavelength absorbing at All4261GAF2(C81L) domain was achieved successfully to the long-wavelength absorbing at the ApcE(1-240/Δ87-130) domain.
School of Life Science ＆ Technology, Hubei University for Nationalities, Enshi 445000, China; Key Laboratory of Biologic Resources Protection and Utilization of Hubei Province, Hubei University for Nationalities, Enshi 445000, China. Electronic address: maqiong110@126.com.
Qiong
Q
Ma
State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, China.
Nan
N
Zhou
State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, China.
Ming
M
Zhou
D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D015870:Gene Expression; D046937:Nostoc / Q000235:genetics*; D052979:Phycobiliproteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
D052979:Phycobiliproteins; D011993:Recombinant Fusion Proteins
Chromophore; Co-expressed; Energy transfer; Fusionprotein; Spectra