Crystal structure of CrSBPase.

(A) Topology is displayed as cartoon with main chain coloured from blue (amino-terminus) to red (carboxy-terminus). (B) Rotated view of A. by x-axis over -90°. (C) Putative active site residues inferred from alignment with FBPase bound to FBP (5l0a, human muscle fructose-1,6-bisphosphatase E69Q mutant in active R-state in complex with fructose-1,6-bisphosphate) are represented in sticks. (D) Surface representation in the B. orientation with putative active site residues coloured in cyan. Water molecule H2O 401 oxygen is represented as a red sphere. (E) Cysteines site chains are represented in spheres. (F) Threonines and serines side chains reported to be the target of phosphorylations are represented in spheres.

Functional characterization of CrSBPase in vitro.

Reported enzymatic activity of CrSBPase assayed for (A) reduction by dithiothreitol (DTT), (B) Magnesium sulfate (MgSO4), and (C) recombinant thioredoxin f2 from Chlamydomonas reinhardtii (CrTRX-f2).

Crystallographic structures of CrSBPase under reducing treatment.

(A-I) Aligned structures of CrSBPase protomers without redox treatment (A, 7b2o chain A) or in the presence of 10 mmol.L−1 TCEP reducing agent (B-I, 7zuv chains A-H).

Molecular dynamics simulation of CrSBPase after reduction.

(A) Overlap of the crystallographic structure of oxidized SBPase and representative structures of equilibrated reduced SBPase during molecular dynamics simulation 2 (MD2). For structures extracted from MD, only residues 109 to 148 are displayed since most of the other residues are closely overlapping those of the crystallographic structure. (B) Overlap of the crystallographic structure of reduced SBPase and representative structures of equilibrated reduced SBPase during MD2. For structures extracted from MD, only residues 109 to 148 are displayed.

Oligomeric structure of CrSBPase.

(A) Asymmetric unit dimer of untreated CrSBPase (7b2o). Chains are represented in cartoon and coloured cyan (chain B) and salmon (chain E). (B) Asymmetric unit homotetramer under reducing treatment (7zuv). Chains A (cyan), C (magenta), F (white), and H (orange) belong to the same asymmetric unit. (C) Close-up view on figure A homodimer interface. Loop 113-ASCAGTAC-120 from chain B (in cyan) is in 5-Å distance of neighbouring chain E (in salmon). C115 and C120 are bonded by a disulfide bridge. (D) Surface representation of 7b2o homodimer, coloured according to conservation score from teal (lowest conservation) to purple (highest conservation) among 150 aligned homologs.

Crystallographic data collections and models building statistics.

Primers used for point mutagenesis.