Allosteric serine hydroxymethyltransferase from monkey liver: Correlation of conformational changes caused by denaturants with the alterations in catalytic activity

1981 ◽  
Vol 3 (2) ◽  
pp. 167-178 ◽  
Author(s):  
Kashi S. Ramesh ◽  
V. S. Anantanarayanan ◽  
N. Appaji Rao
Keyword(s):  
Catalytic Activity ◽  
Conformational Changes ◽  
Serine Hydroxymethyltransferase ◽  
Monkey Liver

scholarly journals Activation of chicken liver dihydrofolate reductase by urea and guanidine hydrochloride is accompanied by conformational change at the active site

1996 ◽  
Vol 315 (1) ◽  
pp. 97-102 ◽  
Author(s):  
Ying-xin FAN ◽  
Ming JU ◽  
Jun-mei ZHOU ◽  
Chen-lu TSOU
Keyword(s):  
Catalytic Activity ◽  
Dihydrofolate Reductase ◽  
Conformational Changes ◽  
Active Site ◽  
Guanidine Hydrochloride ◽  
Chicken Liver ◽  
Peptide Fragments ◽  
Enhanced Fluorescence ◽  
Compact Structure ◽  
Mouse Leukaemia

It has been reported that the activation of dihydrofolate reductase (DHFR) from L1210 mouse leukaemia cells by KCl or thiol modifiers is accompanied by increased digestibility by proteinases [Duffy, Beckman, Peterson, Vitols and Huennekens (1987) J. Biol. Chem. 262, 7028–7033], suggesting a loosening up of the general compact structure of the enzyme. In the present study, the peptide fragments liberated from the chicken liver enzyme by digestion with trypsin in dilute solutions of urea or guanidine hydrochloride (GuHCl) have been separated by FPLC and sequenced. The sequences obtained are unique when compared with the known sequence of DHFR and thus allow the points of proteolytic cleavage identified for the urea- and GuHCl-activated enzyme to be at or near the active site. It was also indicated by the enhanced fluorescence of 2-p-toluidinylnaphthalene 6-sulphonate that conformational changes at the active site in dilute GuHCl parallel GuHCl activation. The above results indicate that the activation of DHFR in dilute denaturants is accompanied by a loosening up of its compact structure especially at or near the active site, suggesting that the flexibility at its active site is essential for the full expression of its catalytic activity.

Lipid fluidity and membrane protein dynamics

1987 ◽  
Vol 7 (11) ◽  
pp. 823-837 ◽  
Author(s):  
Giorgio Lenaz
Keyword(s):  
Electron Transfer ◽  
Catalytic Activity ◽  
Membrane Proteins ◽  
Conformational Changes ◽  
Inner Core ◽  
Lateral Diffusion ◽  
Protein Molecule ◽  
Thermal Fluctuations ◽  
Membrane Diffusion ◽  
Lipid Fluidity

Membrane fluidity plays an important role in cellular functions. Membrane proteins are mobile in the lipid fluid environment; lateral diffusion of membrane proteins is slower than expected by theory, due to both the effect of protein crowding in the membrane and to constraints from the aqueous matrix. A major aspect of diffusion is in macromolecular associations: reduction of dimensionality for membrane diffusion facilitates collisional encounters, as those concerned with receptor-mediated signal transduction and with electron transfer chains. In mitochondrial electron transfer, diffusional control is prevented by the excess of collisional encounters between fast-diffusing ubiquinone and the respiratory complexes. Another aspect of dynamics of membrane proteins is their conformational flexibility. Lipids may induce the optimal conformation for catalytic activity. Breaks in Arrhenius plots of membrane-bound enzymes may be related to lipid fluidity: the break could occur when a limiting viscosity is reached for catalytic activity. Viscosity can affect protein conformational changes by inhibiting thermal fluctuations to the inner core of the protein molecule.

scholarly journals The interaction of troponin C with phosphofructokinase. Comparison with calmodulin

1991 ◽  
Vol 274 (2) ◽  
pp. 445-451 ◽  
Author(s):  
J Lan ◽  
R F Steiner
Keyword(s):  
Catalytic Activity ◽  
Light Scattering ◽  
Dissociation Constant ◽  
Conformational Change ◽  
Conformational Changes ◽  
Troponin C ◽  
Binary Complex ◽  
Fluorescent Labels ◽  
Apparent Dissociation Constant ◽  
Intact Molecule

Phosphofructokinase (PFK) is a calmodulin (CaM)-binding protein [Mayr & Heilmeyer (1983) FEBS Lett. 195, 51-57]. We found that troponin C (TnC), which is homologous to CaM, also binds PFK and affects PFK's catalytic activity, aggregation states and conformational changes as CaM does in most cases. PFK titration of N-acetylaminoethyl-5-naphthylamido-1-sulphonate (‘AEDANS’)-TnC showed that its apparent dissociation constant is comparable with that of PFK-CaM. Fluorescent labels were also used to probe contact regions on TnC and CaM. It is likely that the C-terminal end of the connecting strand of the TnC molecule is close to PFK in the binary complex. Hydrophobic regions of TnC and CaM also possibly play roles in the binding and polymerization of PFK. TnC and CaM deactivate PFK through accelerating PFK conformational change as well as through accelerating PFK tetramer dissociation, as implied in the results of activity, light-scattering, fluorescence and c.d. experiments. The intact molecule of CaM appears to be required to deactivate PFK, because neither half of the CaM molecule has an effect on PFK activity.

scholarly journals Regulation of monkey liver serine hydroxymethyltransferase by nicotinamide nucleotide

1978 ◽  
Vol 174 (3) ◽  
pp. 1055-1058 ◽  
Author(s):  
K S Ramesh ◽  
N A Rao
Keyword(s):  
Serine Hydroxymethyltransferase ◽  
Monkey Liver ◽  
First Time ◽  
Metabolic Interconversion ◽  
Time Lead

The positive homotropic binding of tetrahydrofolate to monkey liver serine hydroxymethyltransferase was abolished on preincubating the enzyme with NADH and NADPH. NAD+ was a negative heterotropic effector, whereas NADP+ was without effect. The allosteric effects of nicotinamide nucleotides on the serine hydroxymethyltransferase, reported for the first time, lead to a better understanding of the regulation of the metabolic interconversion of folate coenzymes.

Mass concentration and activity concentration of creatine kinase isoenzyme MB compared in serum after acute myocardial infarction

1990 ◽  
Vol 36 (1) ◽  
pp. 149-153 ◽  
Author(s):  
J R Delanghe ◽  
A M De Mol ◽  
M L De Buyzere ◽  
I K De Scheerder ◽  
R J Wieme
Keyword(s):  
Myocardial Infarction ◽  
Acute Myocardial Infarction ◽  
Creatine Kinase ◽  
Catalytic Activity ◽  
Conformational Changes ◽  
Mass Concentration ◽  
Activity Concentration ◽  
Steady Increase ◽  
Reference Limit ◽  
Upper Reference Limit

Abstract We compared three current methods (immunoinhibition, "Isomune-CK" immunoprecipitation, and the Tandem-E CKMB II immunoenzymometric assay) for determination of creatine kinase (CK; EC 2.7.3.2) isoenzyme MB in serum. Although results inter-correlated well, the immunoinhibition assay gave higher activity values. Atypical CK forms did not interfere with the immunoprecipitation and immunoenzymometric methods. In acute myocardial infarction the catalytic properties of CK decreased with the enzyme's age, as reflected by a steady increase in activation energy of the catalyzed reaction. In septicemia patients with very low CK and CK-MB catalytic activity, mean CK-MB mass concentration exceeded the upper reference limit, suggesting an increased rate of loss of activity concentration in these patients' sera. Because of the assay's lesser susceptibility to conformational changes at the active site of the enzyme, we suggest that measurement of CK-MB mass concentration is better suited for infarct sizing than measurement of catalytic activity.

scholarly journals Synergistic effects of functionally distinct substitutions in β-lactamase variants shed light on the evolution of bacterial drug resistance

2018 ◽  
Vol 293 (46) ◽  
pp. 17971-17984 ◽  
Author(s):  
Meha P. Patel ◽  
Liya Hu ◽  
Cameron A. Brown ◽  
Zhizeng Sun ◽  
Carolyn J. Adamski ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Conformational Changes ◽  
Active Site ◽  
Enzyme Stability ◽  
Synergistic Effects ◽  
Catalytic Efficiency ◽  
Bacterial Drug Resistance ◽  
Natural Variant ◽  
Loop Conformation ◽  
Shed Light

The CTX-M β-lactamases have emerged as the most widespread extended-spectrum β-lactamases (ESBLs) in Gram-negative bacteria. These enzymes rapidly hydrolyze cefotaxime, but not the related cephalosporin, ceftazidime. ESBL variants have evolved, however, that provide enhanced ceftazidime resistance. We show here that a natural variant at a nonactive site, i.e. second-shell residue N106S, enhances enzyme stability but reduces catalytic efficiency for cefotaxime and ceftazidime and decreases resistance levels. However, when the N106S variant was combined with an active-site variant, D240G, that enhances enzyme catalytic efficiency, but decreases stability, the resultant double mutant exhibited higher resistance levels than predicted on the basis of the phenotypes of each variant. We found that this epistasis is due to compensatory effects, whereby increased stability provided by N106S overrides its cost of decreased catalytic activity. X-ray structures of the variant enzymes in complex with cefotaxime revealed conformational changes in the active-site loop spanning residues 103–106 that were caused by the N106S substitution and relieve steric strain to stabilize the enzyme, but also alter contacts with cefotaxime and thereby reduce catalytic activity. We noted that the 103–106 loop conformation in the N106S-containing variants is different from that of WT CTX-M but nearly identical to that of the non-ESBL, TEM-1 β-lactamase, having a serine at the 106 position. Therefore, residue 106 may serve as a “switch” that toggles the conformations of the 103–106 loop. When it is serine, the loop is in the non-ESBL, TEM-like conformation, and when it is asparagine, the loop is in a CTX-M–like, cefotaximase-favorable conformation.

scholarly journals Voltage-dependent motion of the catalytic region of voltage-sensing phosphatase monitored by a fluorescent amino acid

2016 ◽  
Vol 113 (27) ◽  
pp. 7521-7526 ◽  
Author(s):  
Souhei Sakata ◽  
Yuka Jinno ◽  
Akira Kawanabe ◽  
Yasushi Okamura
Keyword(s):  
Plasma Membrane ◽  
Catalytic Activity ◽  
Amino Acid ◽  
Conformational Changes ◽  
Unnatural Amino Acid ◽  
Voltage Sensor ◽  
Voltage Sensing ◽  
Cytoplasmic Region ◽  
Voltage Dependent ◽  
Voltage Gated Ion Channels

The cytoplasmic region of voltage-sensing phosphatase (VSP) derives the voltage dependence of its catalytic activity from coupling to a voltage sensor homologous to that of voltage-gated ion channels. To assess the conformational changes in the cytoplasmic region upon activation of the voltage sensor, we genetically incorporated a fluorescent unnatural amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap), into the catalytic region of Ciona intestinalis VSP (Ci-VSP). Measurements of Anap fluorescence under voltage clamp in Xenopus oocytes revealed that the catalytic region assumes distinct conformations dependent on the degree of voltage-sensor activation. FRET analysis showed that the catalytic region remains situated beneath the plasma membrane, irrespective of the voltage level. Moreover, Anap fluorescence from a membrane-facing loop in the C2 domain showed a pattern reflecting substrate turnover. These results indicate that the voltage sensor regulates Ci-VSP catalytic activity by causing conformational changes in the entire catalytic region, without changing their distance from the plasma membrane.

The role of conserved residues in the catalytic activity of NDM-1: an approach involving site directed mutagenesis and molecular dynamics

2019 ◽  
Vol 21 (32) ◽  
pp. 17821-17835 ◽  
Author(s):  
Abid Ali ◽  
Rakesh Kumar ◽  
Mir Asif Iquebal ◽  
Sarika Jaiswal ◽  
Dinesh Kumar ◽  
...  
Keyword(s):  
Molecular Dynamics ◽  
Catalytic Activity ◽  
Cell Survival ◽  
Conformational Changes ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Loss Of Function ◽  
Conserved Residues

Drug degraded by enzyme and hence not targeted on to the cell leading to cell survival. After mutation leading to conformational changes and loss of function hence drug was not degraded and remained available for the target to lyse the cell.

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