Triosephosphate Isomerase Requires a Positively Charged Active Site: The Role of Lysine-12

Biochemistry ◽  
1994 ◽  
Vol 33 (10) ◽  
pp. 2809-2814 ◽  
Author(s):  
Patricia J. Lodi ◽  
Louise C. Chang ◽  
Jeremy R. Knowles ◽  
Elizabeth A. Komives
Keyword(s):  
Active Site ◽  
Triosephosphate Isomerase ◽  
Positively Charged

scholarly journals Uncovering the Role of Key Active Site Side Chains in Catalysis: An Extended Brønsted Relationship for Substrate Deprotonation Catalysed by Wild-Type and Variants of Triosephosphate Isomerase

2019 ◽  
Author(s):  
Yashraj S. Kulkarni ◽  
Tina L. Amyes ◽  
John Richard ◽  
Shina Caroline Lynn Kamerlin
Keyword(s):  
Active Site ◽  
Triosephosphate Isomerase ◽  
Valence Bond ◽  
Side Chains ◽  
Wild Type ◽  
Empirical Valence Bond

Manuscript and supporting information outlining an analysis of an extended Brønsted relationship obtained from empirical valence bond simulations of substrate deprotonation catalyzed by wild-type and mutant variants of triosephosphate isomerase.

Functional Role of the Conserved Active Site Proline of Triosephosphate Isomerase†,‡

Biochemistry ◽  
2006 ◽  
Vol 45 (51) ◽  
pp. 15483-15494 ◽  
Author(s):  
Marco G. Casteleijn ◽  
Markus Alahuhta ◽  
Katrin Groebel ◽  
Ibrahim El-Sayed ◽  
Koen Augustyns ◽  
...  
Keyword(s):  
Active Site ◽  
Functional Role ◽  
Triosephosphate Isomerase

scholarly journals Uncovering the Role of Key Active Site Side Chains in Catalysis: An Extended Brønsted Relationship for Substrate Deprotonation Catalysed by Wild-Type and Variants of Triosephosphate Isomerase

2019 ◽  
Author(s):  
Yashraj S. Kulkarni ◽  
Tina L. Amyes ◽  
John Richard ◽  
Shina Caroline Lynn Kamerlin
Keyword(s):  
Active Site ◽  
Triosephosphate Isomerase ◽  
Valence Bond ◽  
Side Chains ◽  
Wild Type ◽  
Empirical Valence Bond

Manuscript and supporting information outlining an analysis of an extended Brønsted relationship obtained from empirical valence bond simulations of substrate deprotonation catalyzed by wild-type and mutant variants of triosephosphate isomerase.

Uncovering the Role of Key Active Site Side Chains in Catalysis: An Extended Brønsted Relationship for Substrate Deprotonation Catalysed by Wild-Type and Variants of Triosephosphate Isomerase

2019 ◽  
Author(s):  
Yashraj S. Kulkarni ◽  
Tina L. Amyes ◽  
John Richard ◽  
Shina Caroline Lynn Kamerlin
Keyword(s):  
Active Site ◽  
Triosephosphate Isomerase ◽  
Valence Bond ◽  
Side Chains ◽  
Wild Type ◽  
Empirical Valence Bond

Manuscript and supporting information outlining an analysis of an extended Brønsted relationship obtained from empirical valence bond simulations of substrate deprotonation catalyzed by wild-type and mutant variants of triosephosphate isomerase.

scholarly journals Structural studies of duck δ2 crystallin mutants provide insight into the role of Thr161 and the 280s loop in catalysis

2004 ◽  
Vol 384 (2) ◽  
pp. 437-447 ◽  
Author(s):  
Liliana M. SAMPALEANU ◽  
Penelope W. CODDING ◽  
Yuri D. LOBSANOV ◽  
May TSAI ◽  
G. David SMITH ◽  
...  
Keyword(s):  
Active Site ◽  
Side Chain ◽  
Structural Studies ◽  
Cycle Enzyme ◽  
Substrate Complex ◽  
Enzyme Substrate ◽  
Catalytically Active ◽  
Urea Cycle Enzyme ◽  
Positively Charged

δ Crystallin, a taxon-specific crystallin present in avian eye lenses, is homologous to the urea cycle enzyme ASL (argininosuccinate lyase). Although there are two δ crystallin isoforms in duck lenses, dδc1 (duck δ1 crystallin) and dδc2 (duck δ2 crystallin), only dδc2 is catalytically active. Previous structural studies have suggested that residues Ser283 and His162 in the multi-subunit active site of dδc2/ASL are the putative catalytic acid/base, while the highly conserved, positively charged Lys289 is thought to help stabilize the carbanion intermediate. The strict conservation of a small hydroxy-containing residue (Thr or Ser) at position 161 adjacent to the putative catalytic base, as well as its proximity to the substrate in the S283A dδc2 enzyme–substrate complex, prompted us to investigate further the role this residue. Structures of the active T161S and inactive T161D dδc2 mutants, as well as T161D complexed with argininosuccinate, have been determined to 2.0 Å resolution. The structures suggest that a hydroxy group is required at position 161 to help correctly position the side chain of Lys289 and the fumarate moiety of the substrate. Threonine is probably favoured over serine, because the interaction of its methyl group with Leu206 would restrict its conformational flexibility. Residues larger than Thr or Ser interfere with substrate binding, supporting previous suggestions that correct positioning of the substrate's fumarate moiety is essential for catalysis to occur. The presence of the 280s loop (i.e. a loop formed by residues 270–290) in the ‘open’ conformation suggests that loop closure, thought to be essential for sequestration of the substrate, may be triggered by the formation of the carbanion or aci-carboxylate intermediates, whose charge distribution more closely mimics that of the sulphate ion found in the active-site region of the inactive dδc1. The 280s loop in dδc1 is in the closed conformation.

Structures of c-di-GMP/cGAMP degrading phosphodiesterase VcEAL: identification of a novel conformational switch and its implication

2019 ◽  
Vol 476 (21) ◽  
pp. 3333-3353 ◽  
Author(s):  
Malti Yadav ◽  
Kamalendu Pal ◽  
Udayaditya Sen
Keyword(s):  
Active Site ◽  
Metal Binding ◽  
Metal Ion ◽  
Triosephosphate Isomerase ◽  
Catalytic Mechanism ◽  
Degradation Mechanism ◽  
Structural Basis ◽  
Conserved Residues ◽  
Phosphodiester Bond ◽  
Cyclic Dinucleotides

Cyclic dinucleotides (CDNs) have emerged as the central molecules that aid bacteria to adapt and thrive in changing environmental conditions. Therefore, tight regulation of intracellular CDN concentration by counteracting the action of dinucleotide cyclases and phosphodiesterases (PDEs) is critical. Here, we demonstrate that a putative stand-alone EAL domain PDE from Vibrio cholerae (VcEAL) is capable to degrade both the second messenger c-di-GMP and hybrid 3′3′-cyclic GMP–AMP (cGAMP). To unveil their degradation mechanism, we have determined high-resolution crystal structures of VcEAL with Ca2+, c-di-GMP-Ca2+, 5′-pGpG-Ca2+ and cGAMP-Ca2+, the latter provides the first structural basis of cGAMP hydrolysis. Structural studies reveal a typical triosephosphate isomerase barrel-fold with substrate c-di-GMP/cGAMP bound in an extended conformation. Highly conserved residues specifically bind the guanine base of c-di-GMP/cGAMP in the G2 site while the semi-conserved nature of residues at the G1 site could act as a specificity determinant. Two metal ions, co-ordinated with six stubbornly conserved residues and two non-bridging scissile phosphate oxygens of c-di-GMP/cGAMP, activate a water molecule for an in-line attack on the phosphodiester bond, supporting two-metal ion-based catalytic mechanism. PDE activity and biofilm assays of several prudently designed mutants collectively demonstrate that VcEAL active site is charge and size optimized. Intriguingly, in VcEAL-5′-pGpG-Ca2+ structure, β5–α5 loop adopts a novel conformation that along with conserved E131 creates a new metal-binding site. This novel conformation along with several subtle changes in the active site designate VcEAL-5′-pGpG-Ca2+ structure quite different from other 5′-pGpG bound structures reported earlier.

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