Photolabeling of the phosphate binding site of mitochondrial F1-ATPase by [32P]azidonitrophenyl phosphate. Identification of the photolabeled amino acid residues

Biochemistry ◽  
1989 ◽  
Vol 28 (4) ◽  
pp. 1442-1448 ◽  
Author(s):  
Jerome Garin ◽  
Laurent Michel ◽  
Alain Dupuis ◽  
Jean Paul Issartel ◽  
Joel Lunardi ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Amino Acid Residues ◽  
Phosphate Binding ◽  
F1 Atpase

Schistosomal Sulfotransferase Interaction with Oxamniquine Involves Hybrid Mechanism of Induced-fit and Conformational Selection

2020 ◽  
Vol 16 (4) ◽  
pp. 451-459 ◽  
Author(s):  
Fortunatus C. Ezebuo ◽  
Ikemefuna C. Uzochukwu
Keyword(s):  
Molecular Dynamics ◽  
Amino Acid ◽  
Binding Energy ◽  
Binding Site ◽  
Conformational Dynamics ◽  
Side Chain ◽  
Amino Acid Residues ◽  
Conformational Selection ◽  
Hybrid Mechanism ◽  
Dynamics Simulations

Background: Sulfotransferase family comprises key enzymes involved in drug metabolism. Oxamniquine is a pro-drug converted into its active form by schistosomal sulfotransferase. The conformational dynamics of side-chain amino acid residues at the binding site of schistosomal sulfotransferase towards activation of oxamniquine has not received attention. Objective: The study investigated the conformational dynamics of binding site residues in free and oxamniquine bound schistosomal sulfotransferase systems and their contribution to the mechanism of oxamniquine activation by schistosomal sulfotransferase using molecular dynamics simulations and binding energy calculations. Methods: Schistosomal sulfotransferase was obtained from Protein Data Bank and both the free and oxamniquine bound forms were subjected to molecular dynamics simulations using GROMACS-4.5.5 after modeling it’s missing amino acid residues with SWISS-MODEL. Amino acid residues at its binding site for oxamniquine was determined and used for Principal Component Analysis and calculations of side-chain dihedrals. In addition, binding energy of the oxamniquine bound system was calculated using g_MMPBSA. Results: The results showed that binding site amino acid residues in free and oxamniquine bound sulfotransferase sampled different conformational space involving several rotameric states. Importantly, Phe45, Ile145 and Leu241 generated newly induced conformations, whereas Phe41 exhibited shift in equilibrium of its conformational distribution. In addition, the result showed binding energy of -130.091 ± 8.800 KJ/mol and Phe45 contributed -9.8576 KJ/mol. Conclusion: The results showed that schistosomal sulfotransferase binds oxamniquine by relying on hybrid mechanism of induced fit and conformational selection models. The findings offer new insight into sulfotransferase engineering and design of new drugs that target sulfotransferase.

scholarly journals Mitigating the Adverse Effects of Polychlorinated Biphenyl Derivatives on Estrogenic Activity via Molecular Modification Techniques

2021 ◽  
Vol 18 (9) ◽  
pp. 4999
Author(s):  
Wei He ◽  
Wenhui Zhang ◽  
Zhenhua Chu ◽  
Yu Li
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Hydrophobic Interaction ◽  
Oestrogen Receptor ◽  
Polychlorinated Biphenyl ◽  
Hydrophobic Interactions ◽  
Estrogenic Activity ◽  
Average Distance ◽  
Amino Acid Residues ◽  
Hydrophobic Amino Acid

The aim of this paper is to explore the mechanism of the change in oestrogenic activity of PCBs molecules before and after modification by designing new PCBs derivatives in combination with molecular docking techniques through the constructed model of oestrogenic activity of PCBs molecules. We found that the weakened hydrophobic interaction between the hydrophobic amino acid residues and hydrophobic substituents at the binding site of PCB derivatives and human oestrogen receptor alpha (hERα) was the main reason for the weakened binding force and reduced anti-oestrogenic activity. It was consistent with the information that the hydrophobic field displayed by the 3D contour maps in the constructed oestrogen activity CoMSIA model was one of the main influencing force fields. The hydrophobic interaction between PCB derivatives and oestrogen-active receptors was negatively correlated with the average distance between hydrophobic substituents and hydrophobic amino acid residues at the hERα-binding site, and positively correlated with the number of hydrophobic amino acid residues. In other words, the smaller the average distance between the hydrophobic amino acid residues at the binding sites between the two and the more the number of them, and the stronger the oestrogen activity expression degree of PCBS derivative molecules. Therefore, hydrophobic interactions between PCB derivatives and the oestrogen receptor can be reduced by altering the microenvironmental conditions in humans. This reduces the ability of PCB derivatives to bind to the oestrogen receptor and can effectively modulate the risk of residual PCB derivatives to produce oestrogenic activity in humans.

Identification of amino acid residues responsible for the α5  subunit binding selectivity of L-655,708, a benzodiazepine binding site ligand at the GABAA receptor

2001 ◽  
Vol 77 (2) ◽  
pp. 445-451 ◽  
Author(s):  
M. Anna Casula ◽  
Frances A. Bromidge ◽  
Gopalan V. Pillai ◽  
Peter B. Wingrove ◽  
Karine Martin ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Gabaa Receptor ◽  
Amino Acid Residues ◽  
Binding Selectivity ◽  
Benzodiazepine Binding

The essential mitotic target of calmodulin is the 110-kilodalton component of the spindle pole body in Saccharomyces cerevisiae

1993 ◽  
Vol 13 (12) ◽  
pp. 7913-7924
Author(s):  
J R Geiser ◽  
H A Sundberg ◽  
B H Chang ◽  
E G Muller ◽  
T N Davis
Keyword(s):  
Amino Acid ◽  
Sequence Analysis ◽  
Binding Site ◽  
Hybrid System ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Amino Acid Residues ◽  
Calmodulin Binding ◽  
Pole Body ◽  
Two Hybrid

Two independent methods identified the spindle pole body component Nuf1p/Spc110p as the essential mitotic target of calmodulin. Extragenic suppressors of cmd1-1 were isolated and found to define three loci, XCM1, XCM2, and XCM3 (extragenic suppressor of cmd1-1). The gene encoding a dominant suppressor allele of XCM1 was cloned. On the basis of DNA sequence analysis, genetic cosegregation, and mutational analysis, XCM1 was identified as NUF1/SPC110. Independently, a C-terminal portion of Nuf1p/Spc110p, amino acid residues 828 to 944, was isolated as a calmodulin-binding protein by the two-hybrid system. As assayed by the two-hybrid system, Nuf1p/Spc110p interacts with wild-type calmodulin and triple-mutant calmodulins defective in binding Ca2+ but not with two mutant calmodulins that confer a temperature-sensitive phenotype. Deletion analysis by the two-hybrid system mapped the calmodulin-binding site of Nuf1p/Spc110p to amino acid residues 900 to 927. Direct binding between calmodulin and Nuf1p/Spc110p was demonstrated by a modified gel overlay assay. Furthermore, indirect immunofluorescence with fixation procedures known to aid visualization of spindle pole body components localized calmodulin to the spindle pole body. Sequence analysis of five suppressor alleles of NUF1/SPC110 indicated that suppression of cmd1-1 occurs by C-terminal truncation of Nuf1p/Spc110p at amino acid residues 856, 863, or 881, thereby removing the calmodulin-binding site.

Comparison of the Amino Acid Residues in the Sixth Transmembrane Domains Accessible in the Binding-Site Crevices of μ, δ, and κ Opioid Receptors†

Biochemistry ◽  
2001 ◽  
Vol 40 (27) ◽  
pp. 8018-8029 ◽  
Author(s):  
Wei Xu ◽  
Jin Li ◽  
Chongguang Chen ◽  
Peng Huang ◽  
Harel Weinstein ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Opioid Receptors ◽  
Transmembrane Domains ◽  
Amino Acid Residues

scholarly journals Gene Cloning and Molecular Characterization of Lysine Decarboxylase from Selenomonas ruminantiumDelineate Its Evolutionary Relationship to Ornithine Decarboxylases from Eukaryotes

2000 ◽  
Vol 182 (23) ◽  
pp. 6732-6741 ◽  
Author(s):  
Yumiko Takatsuka ◽  
Yoshihiro Yamaguchi ◽  
Minenobu Ono ◽  
Yoshiyuki Kamio
Keyword(s):  
Amino Acid ◽  
Substrate Specificity ◽  
Pyridoxal Phosphate ◽  
Evolutionary Relationship ◽  
Binding Domain ◽  
Sequencing Analysis ◽  
Lysine Decarboxylase ◽  
Amino Acid Residues ◽  
Phosphate Binding ◽  
Link Type

ABSTRACT Lysine decarboxylase (LDC; EC 4.1.1.18 ) from Selenomonas ruminantium comprises two identical monomeric subunits of 43 kDa and has decarboxylating activities toward both l-lysine andl-ornithine with similar Km andVmax values (Y. Takatsuka, M. Onoda, T. Sugiyama, K. Muramoto, T. Tomita, and Y. Kamio, Biosci. Biotechnol. Biochem. 62:1063–1069, 1999). Here, the LDC-encoding gene (ldc) of this bacterium was cloned and characterized. DNA sequencing analysis revealed that the amino acid sequence of S. ruminantium LDC is 35% identical to those of eukaryotic ornithine decarboxylases (ODCs; EC 4.1.1.17 ), including the mouse,Saccharomyces cerevisiae, Neurospora crassa,Trypanosoma brucei, and Caenorhabditis elegansenzymes. In addition, 26 amino acid residues, K69, D88, E94, D134, R154, K169, H197, D233, G235, G236, G237, F238, E274, G276, R277, Y278, K294, Y323, Y331, D332, C360, D361, D364, G387, Y389, and F397 (mouse ODC numbering), all of which are implicated in the formation of the pyridoxal phosphate-binding domain and the substrate-binding domain and in dimer stabilization with the eukaryotic ODCs, were also conserved inS. ruminantium LDC. Computer analysis of the putative secondary structure of S. ruminantium LDC showed that it is approximately 70% identical to that of mouse ODC. We identified five amino acid residues, A44, G45, V46, P54, and S322, within the LDC catalytic domain that confer decarboxylase activities toward bothl-lysine and l-ornithine with a substrate specificity ratio of 0.83 (defined as thek cat/Km ratio obtained with l-ornithine relative to that obtained withl-lysine). We have succeeded in converting S. ruminantium LDC to form with a substrate specificity ratio of 58 (70 times that of wild-type LDC) by constructing a mutant protein, A44V/G45T/V46P/P54D/S322A. In this study, we also showed that G350 is a crucial residue for stabilization of the dimer in S. ruminantium LDC.

An actin-binding site containing a conserved motif of charged amino acid residues is essential for the morphogenic effect of villin

Cell ◽  
1992 ◽  
Vol 70 (1) ◽  
pp. 81-92 ◽  
Author(s):  
Evelyne Friederich ◽  
Katie Vancompernolle ◽  
Christian Huet ◽  
Marc Goethals ◽  
Joëlle Finidori ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Actin Binding ◽  
Amino Acid Residues ◽  
Conserved Motif ◽  
Actin Binding Site

Identification of Amino Acid Residues in the C1 Domain of Human Factor Va2 That Affect Phosphatidylserine-Triggered Cofactor Activity.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1711-1711
Author(s):  
Rinku Majumder ◽  
Mary Ann Quinn-Allen ◽  
Barry R. Lentz ◽  
William H. Kane
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Factor Xa ◽  
Amino Acid Residues ◽  
Wild Type ◽  
Hydrophobic Residues ◽  
Factor Va ◽  
C1 Domain ◽  
Cofactor Activity ◽  
Prothrombin Activation

Abstract Tightly associated factors Va and Xa serve as the essential prothrombin-activating complex whose assembly is triggered by occupancy of phosphatidylserine (PS) regulatory sites on both proteins. Factor Va C2 domain contains a binding site for soluble, short chain PS (C6PS) that includes the indole moieties of Trp2063/Trp2064 at the apex of a loop (“spike-1”) (Srivastava A, Quinn-Allen MA, Kim SW, Kane WH, Lentz BR. Biochemistry, 2001, 40(28): 8246–55). Our recent data show that there is a C6PS site in the factor Va2 C1 domain that serves as a regulatory site for assembly and/or activity of the FVa2-FXa complex (Majumder R, Quinn-Allen MA, Kane WH & Lentz BR. Manuscript in Preparation). This C6PS-binding site also involves aromatic and hydrophobic residues (Tyr1956/Tyr1957) located in a homologous loop whose apex is termed “spike 3”. In order to identify the amino acid residues in the C1 domain that contribute to the PS-mediated cofactor activity of factor Va2, charged and hydrophobic residues predicted to be exposed in FVa2-C1 domain were mutated to alanine in clusters of 1–3 mutations per construct. The resultant 20 mutants (R1880A, D1892A, (K1896,E1899)A, (F1900,L1901,Y1903)A, (E1905,R1907)A, Y1917A, (E1923,K1924)A, (K1941,E1942)A, (K1954,H1955)A, (Y1956,L1957)A, Y1956A, L1957A, K1958A, E1964A, K1980A, D1995A, R2019A, (R2023,R2027)A, R2023A, R2027A,) and factor V wild type were expressed in Cos-7 cells followed by activation with thrombin, partial purification and concentration using HiTrap SP HP columns. The specific activities of all factor Va mutants were greater than 70% of wild type, with concentrations in the 1.5-7μM range. Recently it has been shown that two mutants (Y1956, L1957)A and (R2023,R2027)A showed decreased binding to immobilized PS and a selective decrease in prothrombinase activity on membranes containing 5% PS (Saleh M, Peng W, Quinn-Allen MA, Macedo-Ribeiro S, Fuentes-Prior P, Bode W & Kane WH. Thromb. Haemost.2004, 91:16–27). Here we report the rate of prothrombin activation in the presence of 1 nM factor Xa, 5 nM factorVa2 (mutants and wild type) and 400 mM C6PS. Enhancement of cofactor activity (E) of factor Va-C1 wild type and mutants by C6PS was measured using the following equation ( Zhai X, Srivastava A, Drummond DC, Daleke D and Lentz BR. Biochemistry. 2002, 41: 5675–84): \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \[E=\ (r_{Xa.Va.PL}/r_{Xa.PL})/(r_{Xa.Va}/r_{Xa})\] \end{document} Here, rXa·Va·PL is the rate of prothrombin activation measured as the initial slope of the rate of change of normalized DAPA fluorescence with time by enzyme in the presence of factor Va and lipid, and other terms are defined analogously. The cofactor activities of (Y1956, L1957) A, Y1956A and L1957A were drastically reduced (values are 1.1, 4.2 and 5.1 respectively) relative to the cofactor activity of the wild type factor Va2 (15). The cofactor activities of (R2023, R2027) A, E1964A and (K1954, H1955) A were also reduced but to a lesser extent (values are 8, 10.6 and 12 respectively). We plan to monitor the binding of these mutants to C6PS and to factor Xa in the presence of C6PS in order to determine the role of these mutations on the assembly and activity of prothrombinase. Supported by grants from the NHLBI (HL43106 to W. Kane and HL 072827 to BRL).

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