Rationalization of enzyme enantioselectivity by active-site cubic-space models

2002 ◽  
Vol 80 (6) ◽  
pp. 559-564 ◽  
Author(s):  
Gianluca Ottolina ◽  
Giacomo Carrea
Keyword(s):  
Active Site ◽  
Enzymatic Reactions

Active-site models of enzymes are important tools that help researchers in understanding and predicting the stereochemical outcome of enzymatic reactions. In this article, various active-site models for dehydrogenases, hydrolases, oxygenases, and oxynitrilases have been reviewed.

scholarly journals Chi hotspot control of RecBCD helicase-nuclease by long-range intramolecular signaling

2020 ◽  
Author(s):  
Susan K. Amundsen ◽  
Andrew F. Taylor ◽  
Gerald R. Smith
Keyword(s):  
Active Site ◽  
Recognition Site ◽  
Dna Unwinding ◽  
Helicase Domain ◽  
Enzymatic Reactions ◽  
Deficient Mutant ◽  
Long Distance ◽  
Intimate Contact ◽  
Recombination Hotspots ◽  
Dependent Signal

AbstractRepair of broken DNA by homologous recombination requires coordinated enzymatic reactions to prepare it for interaction with intact DNA. The multiple activities of enterobacterial RecBCD helicase-nuclease are coordinated by Chi recombination hotspots (5’ GCTGGTGG 3’) recognized during DNA unwinding. Chi is recognized in a tunnel in RecC but activates the RecB nuclease, >25 Ǻ away. How the Chi-dependent signal travels this long distance has been unknown. We found a Chi-recognition-deficient mutant in the RecB helicase domain located >45 Ǻ from both the Chi-recognition site and the nuclease active site. This unexpected observation led us to find additional mutations that reduced or eliminated Chi hotspot activity in each subunit and widely scattered throughout RecBCD. Each mutation alters the intimate contact between one or another pair of subunits in the crystal or cryoEM structures of RecBCD bound to DNA. Collectively, these mutations span a path ∼185 Ǻ long from the Chi recognition site to the nuclease active site. We discuss these surprising results in the context of an intramolecular signal transduction accounting for many previous observations.

scholarly journals Modelling the Effects of Ageing Time of Starch on the Enzymatic Activity of Three Amylolytic Enzymes

2012 ◽  
Vol 2012 ◽  
pp. 1-13 ◽  
Author(s):  
Nelson P. Guerra ◽  
Lorenzo Pastrana Castro
Keyword(s):  
Enzymatic Activity ◽  
Conformational Changes ◽  
Active Site ◽  
Ageing Time ◽  
Enzymatic Reactions ◽  
Reaction Products ◽  
Inhibitory Effects ◽  
Amylolytic Enzymes ◽  
Modified Forms

The effect of increasing ageing time (t) of starch on the activity of three amylolytic enzymes (Termamyl, San Super, and BAN) was investigated. Although all the enzymatic reactions follow michaelian kinetics,vmaxdecreased significantly (P<0.05) andKMincreased (although not always significantly) with the increase int. The conformational changes produced in the starch chains as a consequence of the ageing seemed to affect negatively the diffusivity of the starch to the active site of the enzymes and the release of the reaction products to the medium. A similar effect was observed when the enzymatic reactions were carried out with unaged starches supplemented with different concentrations of gelatine [G]. The inhibition in the amylolytic activities was best mathematically described by using three modified forms of the Michaelis-Menten model, which included a term to consider, respectively, the linear, exponential, and hyperbolic inhibitory effects oftand [G].

scholarly journals Chi hotspot control of RecBCD helicase-nuclease by long-range intramolecular signaling

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Susan K. Amundsen ◽  
Andrew F. Taylor ◽  
Gerald R. Smith
Keyword(s):  
Active Site ◽  
Recognition Site ◽  
Dna Unwinding ◽  
Helicase Domain ◽  
Enzymatic Reactions ◽  
Deficient Mutant ◽  
Long Distance ◽  
Intimate Contact ◽  
Recombination Hotspots ◽  
Dependent Signal

Abstract Repair of broken DNA by homologous recombination requires coordinated enzymatic reactions to prepare it for interaction with intact DNA. The multiple activities of enterobacterial RecBCD helicase-nuclease are coordinated by Chi recombination hotspots (5′ GCTGGTGG 3′) recognized during DNA unwinding. Chi is recognized in a tunnel in RecC but activates the RecB nuclease, > 25 Ǻ away. How the Chi-dependent signal travels this long distance has been unknown. We found a Chi hotspot-deficient mutant in the RecB helicase domain located > 45 Ǻ from both the Chi-recognition site and the nuclease active site. This unexpected observation led us to find additional mutations that reduced or eliminated Chi hotspot activity in each subunit and widely scattered throughout RecBCD. Each mutation alters the intimate contact between one or another pair of subunits in crystal or cryoEM structures of RecBCD bound to DNA. Collectively, these mutations span a path about 185 Ǻ long from the Chi recognition site to the nuclease active site. We discuss these surprising results in the context of an intramolecular signal transduction accounting for many previous observations.

scholarly journals Crystal Structure of Bacterial Cystathionine Γ-Lyase in The Cysteine Biosynthesis Pathway of Staphylococcus aureus

Crystals ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 656 ◽  
Author(s):  
Dukwon Lee ◽  
Soyeon Jeong ◽  
Jinsook Ahn ◽  
Nam-Chul Ha ◽  
Ae-Ran Kwon
Keyword(s):  
Crystal Structure ◽  
Staphylococcus Aureus ◽  
Active Site ◽  
Structural Changes ◽  
Enzymatic Reactions ◽  
Biosynthesis Pathway ◽  
Active Site Residues ◽  
Structural Comparison ◽  
General Base ◽  
Bond Network

Many enzymes require pyridoxal 5’-phosphate (PLP) as an essential cofactor and share active site residues in mediating diverse enzymatic reactions. Methionine can be converted into cysteine by cystathionine γ-lyases (CGLs) through a transsulfuration reaction dependent on PLP. In bacteria, MccB, also known as YhrB, exhibits CGL activity that cleaves the C–S bond of cystathionine at the γ position. In this study, we determined the crystal structure of MccB from Staphylococcus aureus in its apo- and PLP-bound forms. The structures of MccB exhibited similar molecular arrangements to those of MetC-mediating β-elimination with the same substrate and further illustrated PLP-induced structural changes in MccB. A structural comparison to MetC revealed a longer distance between the N-1 atom of the pyridine ring of PLP and the Oδ atom of the Asp residue, as well as a wider and more flexible active site environment in MccB. We also found a hydrogen bond network in Ser-water-Ser-Glu near the Schiff base nitrogen atom of the PLP molecule and propose the Ser-water-Ser-Glu motif as a general base for the γ-elimination process. Our study suggests the molecular mechanism for how homologous enzymes that use PLP as a cofactor catalyze different reactions with the same active site residues.

scholarly journals Neutron crystallography of copper amine oxidase reveals keto/enolate interconversion of the quinone cofactor and unusual proton sharing

2020 ◽  
Vol 117 (20) ◽  
pp. 10818-10824 ◽  
Author(s):  
Takeshi Murakawa ◽  
Kazuo Kurihara ◽  
Mitsuo Shoji ◽  
Chie Shibazaki ◽  
Tomoko Sunami ◽  
...  
Keyword(s):  
Active Site ◽  
Amine Oxidase ◽  
Catalytic Reactions ◽  
Enzymatic Reactions ◽  
Histidine Residue ◽  
Protonation State ◽  
Copper Amine Oxidase ◽  
Site Structure ◽  
Active Site Structure ◽  
Topa Quinone

Recent advances in neutron crystallographic studies have provided structural bases for quantum behaviors of protons observed in enzymatic reactions. Thus, we resolved the neutron crystal structure of a bacterial copper (Cu) amine oxidase (CAO), which contains a prosthetic Cu ion and a protein-derived redox cofactor, topa quinone (TPQ). We solved hitherto unknown structures of the active site, including a keto/enolate equilibrium of the cofactor with a nonplanar quinone ring, unusual proton sharing between the cofactor and the catalytic base, and metal-induced deprotonation of a histidine residue that coordinates to the Cu. Our findings show a refined active-site structure that gives detailed information on the protonation state of dissociable groups, such as the quinone cofactor, which are critical for catalytic reactions.

Locating Al in the DABCO catalyst before and after Al extraction

1990 ◽  
Vol 48 (4) ◽  
pp. 265-267
Author(s):  
Kathleen B. Reuter
Keyword(s):  
Active Site ◽  
Inverse Relationship ◽  
Transverse Direction ◽  
Reaction Rate ◽  
Acid Phosphate ◽  
Fracture Surfaces ◽  
Al Content ◽  
Before And After ◽  
Relationship Of

The reaction rate and efficiency of piperazine to 1,4-diazabicyclo-octane (DABCO) depends on the Si/Al ratio of the MFI topology catalysts. The Al was shown to be the active site, however, in the Si/Al range of 30-200 the reaction rate increases as the Si/Al ratio increases. The objective of this work was to determine the location and concentration of Al to explain this inverse relationship of Al content with reaction rate.Two silicalite catalysts in the form of 1/16 inch SiO2/Al2O3 bonded extrudates were examined: catalyst A with a Si/Al of 83; and catalyst B, the acid/phosphate Al extracted form of catalyst A, with a Si/Al of 175. Five extrudates from each catalyst were fractured in the transverse direction and particles were obtained from the fracture surfaces near the center of the extrudate diameter. Particles were also obtained from the outside surfaces of five extrudates.

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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