Chl synthase (ChlG) is an important enzyme of the Chl biosynthetic pathway catalyzing attachment of phytol/geranylgeraniol tail to the chlorophyllide molecule. Here we have investigated the Flag-tagged ChlG (f.ChlG) in a complex with two different high-light inducible proteins (Hlips) HliD and HliC. The f.ChlG-Hlips complex binds a Chl a and three different carotenoids, β-carotene, zeaxanthin and myxoxanthophyll. Application of ultrafast time-resolved absorption spectroscopy performed at room and cryogenic temperatures revealed excited-state dynamics of complex-bound pigments. After excitation of Chl a in the complex, excited Chl a is efficiently quenched by a nearby carotenoid molecule via energy transfer from the Chl a Qy state to the carotenoid S1 state. The kinetic analysis of the spectroscopic data revealed that quenching occurs with a time constant of ~2ps and its efficiency is temperature independent. Even though due to its long conjugation myxoxanthophyll appears to be energetically best suited for role of Chl a quencher, based on comparative analysis and spectroscopic data we propose that β-carotene bound to Hlips acts as the quencher rather than myxoxanthophyll and zeaxanthin, which are bound at the f.ChlG and Hlips interface. The S1 state lifetime of the quencher has been determined to be 13ps at room temperature and 21ps at 77K. These results demonstrate that Hlips act as a conserved functional module that prevents photodamage of protein complexes during photosystem assembly or Chl biosynthesis.
Keywords: Carotenoids; Cyanobacteria; Energy transfer; Femtosecond spectroscopy; High-light inducible proteins; Non-photochemical quenching.
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