scholarly journals Reversible deactivation of β-lactamase by quinacillin. Extent of the conformational change in the isolated transitory complex

1986 ◽  
Vol 237 (3) ◽  
pp. 723-730 ◽  
Author(s):  
K C Persaud ◽  
R H Pain ◽  
R Virden
Keyword(s):  
Relative Viscosity ◽  
Fold Increase ◽  
Order Rate Constant ◽  
Side Chain ◽  
Native State ◽  
Significant Difference ◽  
Unfolded State ◽  
Normal Enzyme ◽  
Partially Unfolded ◽  
Asymmetric Environment

Conditions have been established where the deactivation of the beta-lactamase from Staphylococcus aureus PC1 by the penicillin substrate, quinacillin, is close to complete but fully reversible. The temperature-dependence of the rate of re-activation indicated a half-life of about 170 min for the deactivated state at 0 degrees C. Measurement of the relative viscosity of mixtures of enzyme and quinacillin at 8.4 degrees C ruled out any significant difference in shape or solvation between the deactivated and the normal enzyme. C.d. measurements of the deactivated protein, separated from excess quinacillin, showed that the quinacillin side-chain chromophore was bound in an asymmetric environment. The ellipticity associated with the bound quinacillin chromophore decreased with the same first-order rate constant as that for reappearance of enzyme activity. These findings support the accumulation of a deactivated state that contains bound quinacillin or a derivative. Quinacillin caused a 3-fold increase in the rate of 3H exchange-out (at a rate that was low compared with that for the substantially unfolded or expanded protein). However, there was rapid exchange-out of about 50 3H atoms on addition of 1 M-urea to the deactivated enzyme, whereas the same concentration had no effect on the exchange-out of 3H from native enzyme. The interpretation that quinacillin increases the susceptibility of the native state to unfolding in the presence of urea is supported by the demonstration that SO4(2)- ions decreased the rate and extent of deactivation but had no effect on the rate of re-activation, as predicted from the observation that SO4(2)- ions, in competition with urea, stabilize the native state relative to the partially unfolded state H [Mitchinson & Pain (1985) J. Mol. Biol. 184, 331-342].

Cold Shock Domain of the Human Y-Box Protein YB-1. Backbone Dynamics and Equilibrium between the Native State and a Partially Unfolded State†

Biochemistry ◽  
2004 ◽  
Vol 43 (31) ◽  
pp. 10237-10246 ◽  
Author(s):  
Cathelijne P. A. M. Kloks ◽  
Marco Tessari ◽  
Geerten W. Vuister ◽  
Cornelis W. Hilbers
Keyword(s):  
Cold Shock ◽  
Backbone Dynamics ◽  
Native State ◽  
Unfolded State ◽  
Cold Shock Domain ◽  
Partially Unfolded

Salt as a catalyst in the mitochondria: returning cytochrome c to its native state after it misfolds on the surface of cardiolipin containing membranes

2014 ◽  
Vol 50 (28) ◽  
pp. 3674-3676 ◽  
Author(s):  
Leah A. Pandiscia ◽  
Reinhard Schweitzer-Stenner
Keyword(s):  
Cytochrome C ◽  
Native State ◽  
Unfolded State ◽  
Partially Unfolded

After binding to TOCL/DOPC(20%/80%) liposomes ferricytochrome c remains mostly in its partially unfolded state under folding conditions. The addition of 100 mM NaCl switches it back to the native state.

scholarly journals Spectroscopic Studies on Unfolding Processes of Apo-Neuroglobin Induced by Guanidine Hydrochloride and Urea

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Cui Zhang ◽  
Chaohui Gao ◽  
Jianshuai Mu ◽  
Zhanglei Qiu ◽  
Lianzhi Li
Keyword(s):  
Structural Transition ◽  
Guanidine Hydrochloride ◽  
Fluorescence Emission ◽  
Emission Spectra ◽  
Spectroscopic Studies ◽  
Native State ◽  
Fluorescence Emission Spectra ◽  
Unfolded State ◽  
Partially Unfolded ◽  
Far Ultraviolet

Neuroglobin (Ngb), a recently discovered globin, is predominantly expressed in the brain, retina, and other nerve tissues of vertebrates. The unfolding processes of apo-neuroglobin (apoNgb) induced by guanidine hydrochloride (GdnHCl) and urea were investigated by spectroscopic methods. In the unfolding processes, apoNgb's tertiary structural transition was monitored by the changes of intrinsic fluorescence emission spectra, and its secondary structural transition was measured by the changes of far-ultraviolet circular dichroism (CD) spectra. In addition, 8-anilino-1-naphthalenesulfonic acid (ANS), a hydrophobic cluster binding dye, was also used to monitor the unfolding process of apoNgb and to explore its intermediates. Results showed that GdnHCl-induced unfolding of apoNgb was via a three-state pathway, that is, Native state(N)→ Intermediate state(I)→ Unfolded state(U), during which the intermediate was inferred by an increase in fluorescence intensity and the change of CD value. Gibbs free energy changes are 10.2 kJ·mol−1for the first unfolding transition and 14.0 kJ·mol−1for the second transition. However, urea-induced unfolding of apoNgb only underwent a two-state transition: Native state(N)→ Partially unfolded state(P). The result showed that GdnHCl can efficiently affect the conformational states of apoNgb compared with those of urea. The work will benefit to have an understanding of the unfolding mechanism of apoNgb induced by GdnHCl and urea.

scholarly journals Microsecond sub-domain motions and the folding and misfolding of the mouse prion protein

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Rama Reddy Goluguri ◽  
Sreemantee Sen ◽  
Jayant Udgaonkar
Keyword(s):  
Prion Protein ◽  
Conformational Dynamics ◽  
Correlation Spectroscopy ◽  
Native State ◽  
Fluorescence Correlation ◽  
Stochastic Fluctuations ◽  
Unfolded State ◽  
State Ensemble ◽  
Partially Unfolded ◽  
Domain Motions

Protein aggregation appears to originate from partially unfolded conformations that are sampled through stochastic fluctuations of the native protein. It has been a challenge to characterize these fluctuations, under native like conditions. Here, the conformational dynamics of the full-length (23-231) mouse prion protein were studied under native conditions, using photoinduced electron transfer coupled to fluorescence correlation spectroscopy (PET-FCS). The slowest fluctuations could be associated with the folding of the unfolded state to an intermediate state, by the use of microsecond mixing experiments. The two faster fluctuations observed by PET-FCS, could be attributed to fluctuations within the native state ensemble. The addition of salt, which is known to initiate the aggregation of the protein, resulted in an enhancement in the time scale of fluctuations in the core of the protein. The results indicate the importance of native state dynamics in initiating the aggregation of proteins.

Improved Stability of a Protein Vaccine Through Elimination of a Partially Unfolded State

2004 ◽  
Author(s):  
Colleen A. McHugh ◽  
Ralph F. Tammariello ◽  
Charles B. Millard ◽  
John H. Carra
Keyword(s):  
Protein Vaccine ◽  
Unfolded State ◽  
Improved Stability ◽  
Partially Unfolded

737 Prediction of drop out in an adjunctive light treatment trial in patients with non-seasonal depression and evening chronotype

SLEEP ◽  
2021 ◽  
Vol 44 (Supplement_2) ◽  
pp. A287-A288
Author(s):  
Joey W Chan ◽  
Y K Wing ◽  
S P Lam ◽  
Shirley Xin Li ◽  
J Zhang
Keyword(s):  
Clinical Characteristics ◽  
Red Light ◽  
Bright Light ◽  
Fold Increase ◽  
Light Therapy ◽  
Light Treatment ◽  
Drop Out ◽  
Sleep Diary ◽  
Significant Difference ◽  
Seasonal Depression

Abstract Introduction Drop out during treatment hampers therapeutic effect of interventions. The current study examines the possible predictors of drop out during the five-week light treatment in patients with unipolar non-seasonal depression and evening-chronotype. Methods Baseline characteristics including demographics, sleep diary parameters, light treatment prescribed, and early clinical outcomes changes were compared between the Drop out and Non drop out group. Logistic regression analysis was used to examine predictors for drop out. All data were analyzed in a modified intention to treat analysis with last observation carried forward approach. Results A total of 91 subjects (Female 79%, 46.3 ± 11.8 years old) were included in the analysis. There was no significant difference in the baseline demographic and clinical characteristics between the Drop out and Non drop out group. There was also no significant difference in the improvement of clinical parameters over the first week among the two groups. However, treatment non-adherence (in terms of compliance of less than 80% of prescribed duration) over the first treatment week predicts a five-fold increase in risk of drop out during light therapy. (OR: 5.85, CI: 1.414–24.205, p=0.015) after controlling for potential confounders including age, gender, treatment group, patient expectation, and treatment-emergent adverse events. Conclusion This study found that baseline clinical characteristics including depression severity and improvement of depressive symptoms in the initial week did not differ between the Drop out and Non drop out group. The drop out was also not affected by the type of light (dim red versus bright red light), indirectly supporting dim red light as a valid placebo in bright light therapy trial. Treatment adherence is the early phase of light treatment is an important predictor of drop out. Support (if any):

Validation of a locally derived serum calcium adjustment equation: relationship with total 25(OH) vitamin D and PTH levels

2016 ◽  
Vol 70 (1) ◽  
pp. 69-74 ◽  
Author(s):  
Jonathan Joseph Scargill ◽  
John Mark Guy
Keyword(s):  
Vitamin D ◽  
Fold Increase ◽  
Reference Interval ◽  
Albumin Concentration ◽  
Vitamin D Levels ◽  
Significant Difference ◽  
25 Oh Vitamin D ◽  
New Equation ◽  
The Uk ◽  
Adjustment Equation

AimsMany clinical laboratories in the UK use a standard equation to adjust total calcium for albumin concentration. To assess the validity of this practice, we assessed the effect of the use of a traditional and locally derived calcium adjustment equation on parathyroid hormone (PTH) and 25(OH, hydroxy) vitamin D levels.MethodsSamples requested for calcium and albumin measurement over a 6 month period that met inclusion criteria were used to derive a calcium adjustment equation (n=60 941). The traditional and locally derived calcium adjustment equations were then applied to a second cohort of adult patients that underwent calcium measurement over a 1 year period (n=275 456). Patients were classified as hypocalcaemic, normocalcaemic or hypercalcaemic using a UK Pathology Harmony adjusted calcium reference interval (2.2–2.6 mmol/L).ResultsThe local calcium adjustment equation provided a 7.1-fold reduction in the prevalence of hypocalcaemia. Patients classified as hypocalcaemic using the locally derived equation had statistically significantly lower 25(OH) vitamin D and higher PTH levels. A 2.4-fold increase in the prevalence of hypercalcaemia was also observed using the new equation, but with no significant difference in 25(OH) vitamin D or PTH levels.ConclusionsA locally derived calcium adjustment equation reclassified the calcium status of 61 278 (22%) patients. Patients classified as hypocalcaemic by the locally derived equation had significantly lower 25(OH) vitamin D and significantly higher PTH values, providing evidence that use of this adjustment equation generates adjusted calcium results of greater clinical relevance. This study provides further and novel evidence that individual laboratories should determine local equations for adjusted calcium where possible.

scholarly journals Resveratrol Stimulates Cortisol Biosynthesis by Activating SIRT-Dependent Deacetylation of P450scc

Endocrinology ◽  
2012 ◽  
Vol 153 (7) ◽  
pp. 3258-3268 ◽  
Author(s):  
Donghui Li ◽  
Eric B. Dammer ◽  
Marion B. Sewer
Keyword(s):  
Protein Expression ◽  
Fold Increase ◽  
Pyridine Nucleotide ◽  
Nucleotide Metabolism ◽  
Side Chain ◽  
Mitochondrial Enzyme ◽  
Adrenocortical Cells ◽  
Protein Levels ◽  
Chain Cleavage ◽  
Cleavage Enzyme

In the human adrenal cortex, cortisol is synthesized from cholesterol by members of the cytochrome P450 superfamily and hydroxysteroid dehydrogenases. Both the first and last steps of cortisol biosynthesis occur in mitochondria. Based on our previous findings that activation of ACTH signaling changes the ratio of nicotinamide adenine dinucleotide (NAD) phosphate to reduced NAD phosphate in adrenocortical cells, we hypothesized that pyridine nucleotide metabolism may regulate the activity of the mitochondrial NAD+-dependent sirtuin (SIRT) deacetylases. We show that resveratrol increases the protein expression and half-life of P450 side chain cleavage enzyme (P450scc). The effects of resveratrol on P450scc protein levels and acetylation status are dependent on SIRT3 and SIRT5 expression. Stable overexpression of SIRT3 abrogates the cellular content of acetylated P450scc, concomitant with an increase in P450scc protein expression and cortisol secretion. Mutation of K148 and K149 to alanine stabilizes the expression of P450scc and results in a 1.5-fold increase in pregnenolone biosynthesis. Finally, resveratrol also increases the protein expression of P450 11β, another mitochondrial enzyme required for cortisol biosynthesis. Collectively, this study identifies a role for NAD+-dependent SIRT deacetylase activity in regulating the expression of mitochondrial steroidogenic P450.

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