Reconstitution of the core complex (alpha beta)3APCLC8.9 of the phycobilisome from Mastigocladus laminosus using the Lc8.9 linker polypeptide overexpressed in Escherichia coli.
The apcC gene from Mastigocladus laminosus encodes the linker polypeptide LC8.9 located in the phycobilisome core. A T7 RNA polymerase expression system was used to express the linker polypeptide LC8.9 from M. laminosus in Escherichia coli. The apcC gene product was expressed as an inclusion body which was solubilized in a buffer containing 8M urea. Final purification was achieved by ion exchange chromatography on Fractogel TSK CM 650 (S). In addition, a method for preparative isolation of the LC8.9 linker polypeptide from M. laminosus by reverse phase chromatography is presented. Both LC8.9 isolated from M. laminosus and overexpressed in E. coli were capable of reconstituting the complex (alpha beta)3APCLC8.9. The reconstituted complex was identical to preparations isolated from M. laminosus in terms of polypeptide composition, absorption and fluorescence emission spectroscopy.
Betz
M
M
Institut für Molekularbiologie und Biophysik, Eidgenössische Technische Hochschule, Zürich, Schweiz.
Rüegsegger
U
U
Esteban
A M
AM
Sidler
W A
WA
Zuber
H
H
D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D002852:Chromatography, Ion Exchange; D000458:Cyanobacteria / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D045342:Light-Harvesting Protein Complexes; D008969:Molecular Sequence Data; D010452:Peptide Biosynthesis; D010455:Peptides / Q000737:chemistry / Q000302:isolation & purification; D045524:Phycobilisomes; D010798:Phycocyanin / Q000096:biosynthesis* / Q000235:genetics; D010940:Plant Proteins / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
D001426:Bacterial Proteins; D045342:Light-Harvesting Protein Complexes; D010455:Peptides; D045524:Phycobilisomes; D010940:Plant Proteins; C002497:allophycocyanin; D010798:Phycocyanin