3MHG

Dihydroxyacetone phosphate carbanion intermediate in tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes

Summary for 3MHG

• Entry DOI: 10.2210/pdb3mhg/pdb
• Related: 3MHF
• Descriptor: Tagatose 1,6-diphosphate aldolase 2, 1,3-DIHYDROXYACETONEPHOSPHATE, CALCIUM ION, ... (4 entities in total)
• Functional Keywords: tagatose aldolase class i, beta barrel, streptococcus pyogenes, schiff base, carbanion, lyase
• Biological source: Streptococcus pyogenes serotype M1
• Total number of polymer chains: 4
• Total formula weight: 146917.83

Authors

LowKam, C.,Liotard, B. (deposition date: 2010-04-07, release date: 2010-04-28, Last modification date: 2024-11-20)

Primary citation

LowKam, C.,Liotard, B.,Sygusch, J.
Structure of a class I tagatose-1,6-bisphosphate aldolase: investigation into an apparent loss of stereospecificity.
J.Biol.Chem., 285:21143-21152, 2010

PubMed Abstract: Tagatose-1,6-bisphosphate aldolase from Streptococcus pyogenes is a class I aldolase that exhibits a remarkable lack of chiral discrimination with respect to the configuration of hydroxyl groups at both C3 and C4 positions. The enzyme catalyzes the reversible cleavage of four diastereoisomers (fructose 1,6-bisphosphate (FBP), psicose 1,6-bisphosphate, sorbose 1,6-bisphosphate, and tagatose 1,6-bisphosphate) to dihydroxyacetone phosphate (DHAP) and d-glyceraldehyde 3-phosphate with high catalytic efficiency. To investigate its enzymatic mechanism, high resolution crystal structures were determined of both native enzyme and native enzyme in complex with dihydroxyacetone-P. The electron density map revealed a (alpha/beta)(8) fold in each dimeric subunit. Flash-cooled crystals of native enzyme soaked with dihydroxyacetone phosphate trapped a covalent intermediate with carbanionic character at Lys(205), different from the enamine mesomer bound in stereospecific class I FBP aldolase. Structural analysis indicates extensive active site conservation with respect to class I FBP aldolases, including conserved conformational responses to DHAP binding and conserved stereospecific proton transfer at the DHAP C3 carbon mediated by a proximal water molecule. Exchange reactions with tritiated water and tritium-labeled DHAP at C3 hydrogen were carried out in both solution and crystalline state to assess stereochemical control at C3. The kinetic studies show labeling at both pro-R and pro-S C3 positions of DHAP yet detritiation only at the C3 pro-S-labeled position. Detritiation of the C3 pro-R label was not detected and is consistent with preferential cis-trans isomerism about the C2-C3 bond in the carbanion as the mechanism responsible for C3 epimerization in tagatose-1,6-bisphosphate aldolase.

PubMed: 20427286
DOI: 10.1074/jbc.M109.080358

Experimental method

X-RAY DIFFRACTION (1.92 Å)