As a result, glucose can only be converted to acetyl-CoA via the decarboxylation of the three-carbon molecule pyruvate in heterotrophs. However, the glyoxylate shunt, which allows for the synthesis of four-carbon TCA cycle intermediates from acetyl-CoA, has not been found to be reversible to date. Most central metabolic pathways such as glycolysis, fatty acid synthesis, and the TCA cycle have complementary pathways that run in the reverse direction to allow flexible storage and utilization of resources. Furthermore, this domain motion positions a conserved aspartate residue located in the C-terminal domain in the active site of the adjacent monomer, which may serve as a general acid/base in the catalytic mechanism. Alignment of the structures more » shows that a simple hinge motion centered on the conserved residues Phe268 and Thr269 moves the C-terminal domain by about 30° relative to the rest of the molecule.
Structural alignment with the closely related Rhodobacter sphaeroides malyl-CoA lyase complexed with Mg 2+, oxalate and CoA allows a detailed analysis of the domain motion of the enzyme caused by substrate binding. Here, the 1.56 Å resolution X-ray crystal structure of MCL from Methylobacterium extorquens AM1 with bound Mg 2+is presented. MCL enzymes, which are found in a variety of bacteria, are members of the citrate lyase-like family and are involved in the assimilation of one- and two-carbon compounds. Malyl-CoA lyase (MCL) is an Mg 2+-dependent enzyme that catalyzes the reversible cleavage of (2 S)-4-malyl-CoA to yield acetyl-CoA and glyoxylate. This work provides a strategy to improve carbon fixation efficiency in photosynthetic = , We then implement the pathway in a photosynthetic organism Synechococcus elongates PCC7942, and show that it increases the intracellular acetyl-CoA pool and enhances bicarbonate assimilation by roughly 2-fold.
We first functionally demonstrate the design of the MCG pathway in vitro and in Escherichia coli. This pathway converts a C3 metabolite to two acetyl-CoA by fixation of one additional CO2 equivalent, or assimilates glyoxylate, a photorespiration intermediate, to produce acetyl-CoA without net carbon loss. Here we design a synthetic malyl-CoA-glycerate (MCG) pathway to augment the CBB cycle for efficient acetyl-CoA synthesis. The carbon loss in producing acetyl-CoA from decarboxylation of C3 sugar limits the maximum carbon yield of photosynthesis. The Calvin–Benson–Bassham (CBB) cycle is presumably evolved for optimal synthesis of C3 sugars, but not for the production of C2 metabolite acetyl-CoA.
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