Crystal structure analyses of reconstituted water-soluble chlorophyll proteins from kale

Department of Biomolecular Science, Toho University
○Hiroshi Hara Hiroyuki Satoh Isao Oonishi Akira Uchida

Water-soluble chlorophyll proteins (WSCP) have been considered as a chlorophyll (Chl) carrier in the Chl metabolic pathway because of its water-solubility, stress- inducibility and ability to protect Chl against photooxidation. WSCPs can be classified into two classes based on their photoconvertibility (Classes I and II). WSCPs having no photoconvertibility can be categorized further into two subtypes according to their Chl a/b ratio (Classes IIA and IIB). Class IIA WSCPs exhibit a Chl a/b ratio higher than the ratio in total leaf extract and Class IIB WSCPs possess lower ratio. In order to investigate whether Class II WSCPs discriminate between Chls a and b, we reconstituted kale-WSCP containing solely Chl a or b, and determined the crystal structures.
A cDNA for kale-WSCP was used to express the apo-WSCP in Escherichia coli. A WSCP containing Chl a (WSCP-a) or Chl b (WSCP-b) was reconstituted in vitro. Crystals of WSCPs-a and -b were obtained from similar conditions by hanging-drop vapor-diffusion method. Native data sets of WSCPs-a and -b were collected with synchrotron radiation up to 1.9 Å and 1.6 Å, respectively at 100 K at the KEK-PF. The structures of WSCPs-a and -b were determined by the molecular replacement using a native kale-WSCP as a search model.
Current refined models of WSCP-a and -b have R-factors of 18.4 and 19.1 % and free R-factors of 24.5 and 23.6 %, respecticely. The further refinements are in progress.