One-helix protein 2 is not required for the synthesis of photosystem II subunit D1 in Chlamydomonas.

In land plants and cyanobacteria, co-translational association of chlorophyll (Chl) to the nascent D1 polypeptide, a reaction center protein of photosystem II (PSII), requires a Chl binding complex consisting of a short-chain dehydrogenase (high chlorophyll fluorescence 244 [HCF244]/uncharacterized protein 39 [Ycf39]) and one-helix proteins (OHP1 and OHP2 in chloroplasts) of the light-harvesting antenna complex superfamily. Here, we show that an ohp2 mutant of the green alga Chlamydomonas (Chlamydomonas reinhardtii) fails to accumulate core PSII subunits, in particular D1 (encoded by the psbA mRNA). Extragenic suppressors arose at high frequency, suggesting the existence of another route for Chl association to PSII. The ohp2 mutant was complemented by the Arabidopsis (Arabidopsis thaliana) ortholog. In contrast to land plants, where psbA translation is prevented in the absence of OHP2, ribosome profiling experiments showed that the Chlamydomonas mutant translates the psbA transcript over its full length. Pulse labeling suggested that D1 is degraded during or immediately after translation. The translation of other PSII subunits was affected by assembly-controlled translational regulation. Proteomics showed that HCF244, a translation factor which associates with and is stabilized by OHP2 in land plants, still partly accumulates in the Chlamydomonas ohp2 mutant, explaining the persistence of psbA translation. Several Chl biosynthesis enzymes overaccumulate in the mutant membranes. Partial inactivation of a D1-degrading protease restored a low level of PSII activity in an ohp2 background, but not photoautotrophy. Taken together, our data suggest that OHP2 is not required for psbA translation in Chlamydomonas, but is necessary for D1 stabilization.

Conformation of receptor adopted upon interaction with virus revealed by site-specific fluorescence quenchers and FRET analysis.

Human rhinovirus serotype 2 (HRV2) specifically binds to very-low-density lipoprotein receptor (VLDLR). Among the eight extracellular repeats of VLDLR, the third module (V3) has the highest affinity for the virus, and 12 copies of the genetically engineered concatamer V33333-His(6) were found to bind per virus particle. In the present study, ring formation of V33333-His(6) about each of the 12 5-fold symmetry axes on HRV2 was demonstrated by fluorescence resonance energy transfer (FRET) between donor and acceptor on N- and C-terminus, respectively. In particular, the N-terminus of V33333-His(6) was labeled with fluorescein, and the C-terminus with a new quencher which was bound to the His(6) tag with nanomolar affinity (K(d) approximately 10(-8) M) in the presence of 2 microM NiCl(2).