ABSTRACT
Various plants possess hydrophilic chlorophyll (Chl) proteins known as water-soluble Chl-binding proteins (WSCPs). WSCPs exist in two forms: Class I and Class II, of which Class I alone exhibits unique photoconvertibility. Although numerous genes encoding Class II WSCPs have been identified and the molecular properties of their recombinant proteins have been well characterized, no Class I WSCP gene has been identified to date. In this study, we cloned the cDNA and a gene encoding the Class I WSCP of Chenopodium album (CaWSCP). Sequence analyses revealed that CaWSCP comprises a single exon corresponding to 585bp of an open reading frame encoding 195 amino acid residues. The CaWSCP protein sequence possesses a signature of DUF538, a protein superfamily of unknown function found almost exclusively in Embryophyta. The recombinant CaWSCP was expressed in Escherichia coli as a hexa-histidine fusion protein (CaWSCP-His) that removes Chls from the thylakoid. Under visible light illumination, the reconstituted CaWSCP-His was successfully photoconverted into a different pigment with an absorption spectrum identical to that of native CaWSCP. Interestingly, while CaWSCP-His could bind both Chl a and Chl b, photoconversion occurred only in CaWSCP-His reconstituted with Chl a.
Subject(s)
Chenopodium album/metabolism , Chlorophyll Binding Proteins/genetics , Amino Acid Sequence , Base Sequence , Chlorophyll Binding Proteins/chemistry , Chlorophyll Binding Proteins/metabolism , Cloning, Molecular , Molecular Sequence Data , Sequence Analysis, ProteinABSTRACT
A water-soluble chlorophyll-binding protein (WSCP) with photoconvertibility from Chenopodium album was extracted, purified and crystallized in a darkroom. Green crystals suitable for data collection appeared in about 10 d. A native data set was collected to 2.0 A resolution at 100 K. The space group of the crystal was determined to be orthorhombic I222 or I2(1)2(1)2(1), with unit-cell parameters a = 48.13, b = 60.59, c = 107.21 A. Preliminary analysis of the X-ray data indicated that there is one molecule per asymmetric unit.
