title: Biosynthesis of a stable allophycocyanin beta subunit in metabolically engineered Escherichia coli.
abstract: Allophycocyanin (APC) is widely used as a fluorescent tag for fluorescence detection techniques. In this study, the apcB gene from a thermophilic cyanobacterium strain was cloned and ligated into an expression vector to construct a pathway for the biosynthesis of an allophycocyanin beta subunit (holo-apcBT) in Escherichia coli. Isopropyl β-d-1-thiogalactopyranoside induction successfully reconstituted holo-apcBT in E. coli. The recombinant holo-apcB showed spectroscopic properties similar to native APC. The stability and spectroscopic properties of this protein were then compared with another recombinant allophycocyanin beta subunit (holo-apcBM) whose apcB gene was cloned from mesophilic cyanobacterium. At high temperatures and during the course of storage, holo-apcBT was significantly more stable than holo-apcBM. In addition, holo-apcBT had an unexpectedly higher extinction coefficient and fluorescence quantum yield than holo-apcBM, suggesting that holo-apcBT would be a promising fluorescent tag and serve as a substitute for native APC.
authors.affiliation: Key Laboratory of Experimental Marine Biology, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China.
authors.forename: Huaxin
authors.initials: H
authors.lastname: Chen
authors.forename: Hanzhi
authors.initials: H
authors.lastname: Lin
authors.forename: Fuchao
authors.initials: F
authors.lastname: Li
authors.forename: Peng
authors.initials: P
authors.lastname: Jiang
authors.forename: Song
authors.initials: S
authors.lastname: Qin
mesh_terms: D000595:Amino Acid Sequence; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005453:Fluorescence; D060847:Metabolic Engineering; D008969:Molecular Sequence Data; D010798:Phycocyanin / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D021122:Protein Subunits / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D016415:Sequence Alignment
chemical_list: D021122:Protein Subunits; D011994:Recombinant Proteins; C002497:allophycocyanin; D010798:Phycocyanin