Structures and enzymatic mechanisms of phycobiliprotein lyases CpcE/F and PecE/F

Abstract

The light-harvesting phycobilisome in cyanobacteria and red algae requires the lyase-catalyzed chromophorylation of phycobiliproteins. There are three functionally distinct lyase families known. The heterodimeric E/F type is specific for attaching bilins covalently to α-subunits of phycocyanins and phycoerythrins. Unlike other lyases, the lyase also has chromophore-detaching activity. A subclass of the E/F-type lyases is, furthermore, capable of chemically modifying the chromophore. Although these enzymes were characterized >25 y ago, their structures remained unknown. We determined the crystal structure of the heterodimer of CpcE/F from sp. PCC7120 at 1.89-Å resolution. Both subunits are twisted, crescent-shaped α-solenoid structures. CpcE has 15 and CpcF 10 helices. The inner (concave) layer of CpcE (helices h2, 4, 6, 8, 10, 12, and 14) and the outer (convex) layer of CpcF (h16, 18, 20, 22, and 24) form a cavity into which the phycocyanobilin chromophore can be modeled. This location of the chromophore is supported by mutations at the interface between the subunits and within the cavity. The structure of a structurally related, isomerizing lyase, PecE/F, that converts phycocyanobilin into phycoviolobilin, was modeled using the CpcE/F structure as template. A HC motif critical for the isomerase activity of PecE/F is located at the loop between h20 and h21, supporting the proposal that the nucleophilic addition of Cys-88 to C10 of phycocyanobilin induces the isomerization of phycocyanobilin into phycoviolobilin. Also, the structure of NblB, involved in phycobilisome degradation could be modeled using CpcE as template. Combined with CpcF, NblB shows a low chromophore-detaching activity.

Decomposition of cyanobacterial light harvesting complexes: NblA‐dependent role of the bilin lyase homolog NblB

“…A recent attempt to detect NblB activity in vitro did not reveal phycocyanobilin detachment (Zhao et al ., ), in line with data reported here demonstrating dependence of NblB activity on the presence of NblA.…”

“…A recent study determined the crystal structure of the heterodimer of CpcE/F from Nostoc sp. PCC7120 and used CpcE as a template to model NblB of this cyanobacterium (Zhao et al ., ). Given the sequence conservation between NblB proteins of Nostoc and S .…”

“…This modelling revealed that R26 and W175 point inwards towards a cavity in the structure, whereas W113 points outwards (Figure S2). Of note, modelling of a chromophore into the structure determined for CpcE/F from Nostoc together with a mutational approach supported the suggestion that the chromophore resides in a cavity formed between CpcE and CpcF (Zhao et al ., ). Whether NblB functions in a monomeric form or as a dimer, it is possible that R26 and W175 directly affect the chromophore binding cavity, whereas mutation of W113 elicits a structural change that indirectly affects NblB activity.…”

CpeF is the bilin lyase that ligates the doubly linked phycoerythrobilin on β-phycoerythrin in the cyanobacterium Fremyella diplosiphon

“…We further analyzed the CpeF sequence using the Phyre 2 modeling program (38) to predict the secondary and 3D structures. Using PDB code 5N3U as a template (27), Phyre 2 predicted that the CpeF protein has an ␣-solenoid type structure ( Fig. S1C), consistent with the structure of the only E/Ftype lyase crystallized to date, CpcE/CpcF from Nostoc sp.…”

“…The crystal structure of CpcE/CpcF from Nostoc sp. PCC 7120 was determined, and the structure is primarily ␣-helical consisting of crescent-shaped elongated superhelices or solenoids (27).…”