scholarly journals Dual role of strigolactone receptor signaling partner in inhibiting substrate hydrolysis

2021 ◽  
Author(s):  
Briana L Sobecks ◽  
Jiming Chen ◽  
Diwakar Shukla
Keyword(s):  
Active Site ◽  
Conformational Stability ◽  
Critical Role ◽  
Md Simulations ◽  
Binding Pocket ◽  
Conformational Ensemble ◽  
Downstream Signaling ◽  
F Box Protein ◽  
Substrate Hydrolysis

Plant branch and root growth relies on metabolism of the strigolactone (SL) hormone. The interaction between the SL molecule, Oryza sativa DWARF14 (D14) SL receptor, and D3 F-box protein has been shown to play a critical role in SL perception. Previously, it was believed that D3 only interacts with the closed form of D14 to induce downstream signaling, but recent experiments indicate that D3, as well as its C-terminal helix (CTH), can interact with the open form as well to inhibit strigolactone signaling. Two hypotheses for the CTH induced inhibition are that either the CTH affects the conformational ensemble of D14 by stabilizing catalytically inactive states, or the CTH interacts with SLs in a way that prevents them from entering the binding pocket. In this study, we have performed molecular dynamics (MD) simulations to assess the validity of these hypotheses. We used an apo system with only D14 and the CTH to test the active site conformational stability and a holo system with D14, the CTH, and an SL molecule to test the interaction between the SL and CTH. Our simulations show that the CTH affects both active site conformation and the ability of SLs to move into the binding pocket. In the apo system, the CTH allosterically stabilized catalytic residues into their inactive conformation. In the holo system, significant interactions between SLs and the CTH hindered the ability of SLs to enter the D14 binding pocket. These two mechanisms account for the observed decrease in SL binding to D14 and subsequent ligand hydrolysis in the presence of the CTH.

Unveiling the critical role of active site interaction in single atom catalyst towards hydrogen evolution catalysis

Nano Energy ◽  
2022 ◽  
Vol 93 ◽  
pp. 106819
Author(s):  
Feng Li ◽  
Gao-Feng Han ◽  
Yunfei Bu ◽  
Shanshan Chen ◽  
Ishfaq Ahmad ◽  
...  
Keyword(s):  
Hydrogen Evolution ◽  
Active Site ◽  
Critical Role ◽  
Single Atom

scholarly journals Albino mutants of Streptomyces glaucescens tyrosinase

1991 ◽  
Vol 274 (3) ◽  
pp. 707-713 ◽  
Author(s):  
M P Jackman ◽  
A Hajnal ◽  
K Lerch
Keyword(s):  
Active Site ◽  
Critical Role ◽  
Site Directed Mutagenesis ◽  
Mutant Proteins ◽  
Carbonyl Derivatives ◽  
Hydrogen Bonding Interactions ◽  
Albino Phenotype ◽  
Derivatives Of ◽  
Streptomyces Glaucescens

Site-directed mutagenesis was used to determine the functional role of several residues of Streptomyces glaucescens tyrosinase. Replacement of His-37, -53, -193 or -215 by glutamine yields albino phenotypes, as determined by expression on melanin-indicator plates. The purified mutant proteins display no detectable oxy-enzyme and increased Cu lability at the binuclear active site. The carbonyl derivatives of H189Q and H193Q luminesce, with lambda max. displaced more than 25 nm to a longer wavelength compared with native tyrosinase. The remaining histidine mutants display no detectable luminescence. The results are consistent with these histidine residues (together with His-62 and His-189 reported earlier) acting as Cu ligands in the Streptomyces glaucescens enzyme. Conservative substitution of the invariant Asn-190 by glutamine also gives an albino phenotype, no detectable oxy-enzyme and labilization of active-site Cu. The luminescence spectrum of carbonyl-N190Q, however, closely resembles that of the native enzyme under conditions promoting double Cu occupancy of the catalytic site. A critical role for Asn-190 in active-site hydrogen-bonding interactions is proposed.

scholarly journals Mutational Analysis of VIM-2 Reveals an Essential Determinant for Metallo-β-Lactamase Stability and Folding

2010 ◽  
Vol 54 (8) ◽  
pp. 3197-3204 ◽  
Author(s):  
Luisa Borgianni ◽  
Julie Vandenameele ◽  
André Matagne ◽  
Luca Bini ◽  
Robert A. Bonomo ◽  
...  
Keyword(s):  
Mutational Analysis ◽  
Critical Role ◽  
Kinetic Studies ◽  
Amino Acid Position ◽  
Structural Knowledge ◽  
Saturation Mutagenesis ◽  
Biochemical Analyses ◽  
The Stability ◽  
Substrate Hydrolysis

ABSTRACT Metallo-β-lactamase (MBL)-producing bacteria are emerging worldwide and represent a formidable threat to the efficacy of relevant β-lactams, including carbapenems, expanded-spectrum cephalosporins, and β-lactamase inactivator/β-lactam combinations. VIM-2 is currently the most widespread MBL and represents a primary target for MBL inhibitor research, the clinical need for which is expected to further increase in the future. Using a saturation mutagenesis approach, we probed the importance of four residues (Phe-61, Ala-64, Tyr-67, and Trp-87) located close to the VIM-2 active site and putatively relevant to the enzyme activity based on structural knowledge of the enzyme and on structure-activity relationships of the subclass B1 MBLs. The ampicillin MIC values shown by the various mutants were affected very differently depending on the randomized amino acid position. Position 64 appeared to be rather tolerant to substitution, and kinetic studies showed that the A64W mutation did not significantly affect substrate hydrolysis or binding, representing an important difference from IMP-type enzymes. Phe-61 and Tyr-67 could be replaced with several amino acids without the ampicillin MIC being significantly affected, but in contrast, Trp-87 was found to be critical for ampicillin resistance. Further kinetic and biochemical analyses of W87A and W87F variants showed that this residue is apparently important for the structure and proper folding of the enzyme but, surprisingly, not for its catalytic activity. These data support the critical role of residue 87 in the stability and folding of VIM-2 and might have strong implications for MBL inhibitor design, as this residue would represent an ideal target for interaction with small molecules.

The critical role of dimer formation in monosaccharides binding to human serum albumin

2018 ◽  
Vol 20 (5) ◽  
pp. 3249-3257 ◽  
Author(s):  
Prapasiri Pongprayoon ◽  
Toshifumi Mori
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Critical Role ◽  
Binding Pocket ◽  
Dimer Formation ◽  
Dimeric Structure

Monosaccharides are found to bind tightly to human serum albumin when a dimeric structure is formed in the binding pocket.

Molecular docking and molecular dynamics simulations of fumarate hydratase and its mutant H235N complexed with pyromellitic acid and citrate

2017 ◽  
Vol 15 (06) ◽  
pp. 1750026 ◽  
Author(s):  
S. Subasri ◽  
Santosh Kumar Chaudhary ◽  
K. Sekar ◽  
Manish Kesherwani ◽  
D. Velmurugan
Keyword(s):  
Molecular Dynamics ◽  
Active Site ◽  
Model Building ◽  
Hydrophobic Interactions ◽  
Conformational Stability ◽  
Fumarate Hydratase ◽  
Md Simulations ◽  
Major Effect ◽  
Active Site Residues ◽  
Pyromellitic Acid

Fumarase catalyzes the reversible, stereospecific hydration/dehydration of fumarate to L-malate during the Kreb’s cycle. In the crystal structure of the tetrameric fumarase, it was found that some of the active site residues S145, T147, N188 G364 and H235 had water-mediated hydrogen bonding interactions with pyromellitic acid and citrate which help to the protonation state for the conversion of fumarate to malate. When His 235 is mutated with Asn (H235N), water-mediated interactions were lost due to the shifting of active site water molecule by 0.7 Å away. Molecular dynamics (MD) simulations were also carried out by NAMD and analyzed using Assisted Model Building with Energy Refinement (AMBER) program to better understand the conformational stability and other aspects during the binding of pyromellitic acid and citrate with native and mutant FH. The role of hydrogen bonds and hydrophobic interactions was also analyzed. The present study confirms that the H235N mutation has a major effect on the catalytic activity of fumarase which is evident from the biochemical studies.

scholarly journals MDM2 mediates fibroblast activation and renal tubulointerstitial fibrosis via a p53-independent pathway

2017 ◽  
Vol 312 (4) ◽  
pp. F760-F768 ◽  
Author(s):  
Chen Ye ◽  
Hui Tang ◽  
Zheng Zhao ◽  
Chun-Tao Lei ◽  
Chao-Qun You ◽  
...  
Keyword(s):  
Critical Role ◽  
Tubulointerstitial Fibrosis ◽  
Podocyte Injury ◽  
Fibroblast Activation ◽  
Downstream Signaling ◽  
Renal Tubulointerstitial Fibrosis ◽  
Inflammatory Processes ◽  
Tgf Β1

It is well recognized that murine double minute gene 2 (MDM2) plays a critical role in cell proliferation and inflammatory processes during tumorigenesis. It is also reported that MDM2 is expressed in glomeruli and involved in podocyte injury. However, whether MDM2 is implicated in renal fibrosis remains unclear. Here we investigated the role of MDM2 in tubulointerstitial fibrosis (TIF). By immunohistochemical staining and Western blotting we confirmed that MDM2 is upregulated in the tubulointerstitial compartment in patients with TIF and unilateral urethral obstruction (UUO) mice, which mainly originates from myofibroblasts. Consistently, in vitro MDM2 is increased in TGF-β1-treated fibroblasts, one of the major sources of collagen-producing myofibroblasts during TIF, along with fibroblast activation. Importantly, genetic deletion of MDM2 significantly attenuates fibroblast activation. We then analyzed the possible downstream signaling of MDM2 during fibroblast activation. p53-dependent pathway is the classic downstream signaling of MDM2, and Nutlin-3 is a small molecular inhibitor of MDM2-p53 interaction. To our surprise, Nutlin-3 could not ameliorate fibroblast activation in vitro and TIF in UUO mice. However, we found that Notch1 signaling is attenuated during fibroblast activation, which could be markedly rescued by MDM2 knockdown. Overexpression of intracellular domain of Notch1 (NICD) by plasmid could obviously minimize fibroblast activation induced by TGF-β1. In addition, the degradation of NICD is strikingly suppressed by PYR-41, an inhibitor of ubiquitin-activating enzyme E1, and proteasome inhibitor MG132. Taken together, our findings provide the first evidence that MDM2 is involved in fibroblast activation and TIF, which associates with Notch1 ubiquitination and proteasome degradation.

scholarly journals Correction to “Critical Role of the Secondary Binding Pocket in Modulating the Enzymatic Activity of DUSP5 toward Phosphorylated ERKs”

Biochemistry ◽  
2018 ◽  
Vol 57 (26) ◽  
pp. 3987-3987
Author(s):  
Marat R. Talipov ◽  
Jaladhi Nayak ◽  
Michael Lepley ◽  
Robert D. Bongard ◽  
Daniel S. Sem ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Critical Role ◽  
Binding Pocket

scholarly journals Conformational Landscapes of Halohydrin Dehalogenases and Their Accessible Active Site Tunnels

Catalysts ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 1403
Author(s):  
Miquel Estévez-Gay ◽  
Javier Iglesias-Fernández ◽  
Sílvia Osuna
Keyword(s):  
Active Site ◽  
Ring Opening ◽  
Md Simulations ◽  
Binding Pocket ◽  
Epoxide Ring ◽  
Epoxide Ring Opening ◽  
Breathing Motion ◽  
Reduction Techniques ◽  
Long Time ◽  
Linear Dimensionality Reduction

Halohydrin dehalogenases (HHDH) are industrially relevant biocatalysts exhibiting a promiscuous epoxide-ring opening reactivity in the presence of small nucleophiles, thus giving access to novel carbon–carbon, carbon–oxygen, carbon–nitrogen, and carbon–sulfur bonds. Recently, the repertoire of HHDH has been expanded, providing access to some novel HHDH subclasses exhibiting a broader epoxide substrate scope. In this work, we develop a computational approach based on the application of linear and non-linear dimensionality reduction techniques to long time-scale Molecular Dynamics (MD) simulations to study the HHDH conformational landscapes. We couple the analysis of the conformational landscapes to CAVER calculations to assess their impact on the active site tunnels and potential ability towards bulky epoxide ring opening reaction. Our study indicates that the analyzed HHDHs subclasses share a common breathing motion of the halide binding pocket, but present large deviations in the loops adjacent to the active site pocket and N-terminal regions. Such conformational differences affect the available tunnels for epoxide binding to the active site. The superior activity of the HHDH G subclass towards bulkier substrates is explained by the additional structural elements delimiting the active site region, its rich conformational heterogeneity, and the substantially wider and frequently observed active site tunnels. This study therefore provides key information for HHDH promiscuity and engineering.

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