2X16

Crystallographic binding studies with an engineered monomeric variant of triosephosphate isomerase

Summary for 2X16

• Entry DOI: 10.2210/pdb2x16/pdb
• Related: 1AG1 1DKW 1IIG 1IIH 1KV5 1ML1 1MSS 1MTM 1TPD 1TPE 1TPF 1TRD 1TRI 1TSI 1TTI 1TTJ 2J24 2J27 2V0T 2V2C 2V2D 2V2H 2V5L 2VEI 2VEK 2VEL 2VEM 2VEN 2WSQ 2WSR 2X0M 3TIM 4TIM 5TIM 6TIM
• Descriptor: TRIOSEPHOSPHATE ISOMERASE, GLYCOSOMAL (2 entities in total)
• Functional Keywords: pentose shunt, gluconeogenesis, lipid synthesis, substrate specificity, fatty acid biosynthesis, tim barrel, peroxisome, glycolysis, isomerase, glycosome
• Biological source: TRYPANOSOMA BRUCEI BRUCEI
• Total number of polymer chains: 2
• Total formula weight: 51318.58

Authors

Salin, M.,Kapetaniou, E.G.,Vaismaa, M.,Lajunen, M.,Casteleijn, M.G.,Neubauer, P.,Salmon, L.,Wierenga, R. (deposition date: 2009-12-21, release date: 2009-12-29, Last modification date: 2023-12-20)

Primary citation

Salin, M.,Kapetaniou, E.G.,Vaismaa, M.,Lajunen, M.,Casteleijn, M.G.,Neubauer, P.,Salmon, L.,Wierenga, R.
Crystallographic Binding Studies with an Engineered Monomeric Variant of Triosephosphate Isomerase
Acta Crystallogr.,Sect.D, 66:934-, 2010

PubMed Abstract: Crystallographic binding studies have been carried out to probe the active-site binding properties of a monomeric variant (A-TIM) of triosephosphate isomerase (TIM). These binding studies are part of a structure-based directed-evolution project aimed towards changing the substrate specificity of monomeric TIM and are therefore aimed at finding binders which are substrate-like molecules. A-TIM has a modified more extended binding pocket between loop-7 and loop-8 compared with wild-type TIM. The A-TIM crystals were grown in the presence of citrate, which is bound in the active site of each of the two molecules in the asymmetric unit. In this complex, the active-site loops loop-6 and loop-7 adopt the closed conformation, similar to that observed in liganded wild-type TIM. Extensive crystal-soaking protocols have been developed to flush the bound citrate out of the active-site pocket of both molecules and the crystal structure shows that the unliganded open conformation of the A-TIM active site is the same as in unliganded wild-type TIM. It is also shown that sulfonate compounds corresponding to the transition-state analogue 2-phosphoglycolate bind in the active site, which has a closed conformation. It is also shown that the new binding pocket of A-TIM can bind 3-phosphoglycerate (3PGA; an analogue of a C4-sugar phosphate) and 4-phospho-D-erythronohydroxamic acid (4PEH; an analogue of a C5-sugar phosphate). Therefore, these studies have provided a rationale for starting directed-evolution experiments aimed at generating the catalytic properties of a C5-sugar phosphate isomerase on the A-TIM framework.

PubMed: 20693693
DOI: 10.1107/S0907444910025710

Experimental method

X-RAY DIFFRACTION (2.13 Å)