scholarly journals The effect of 1,4-diaminobutanone on the stability of ornithine decarboxylase from Aspergillus nidulans

1977 ◽  
Vol 166 (3) ◽  
pp. 635-637 ◽  
Author(s):  
L Stevens ◽  
I M McKinnon
Keyword(s):  
Aspergillus Nidulans ◽  
Ornithine Decarboxylase ◽  
Half Life ◽  
Competitive Inhibitor ◽  
The Stability

1,4-Diaminobutanone, a competitive inhibitor of ornithine decarboxylase in Aspergillus nidulans, is able to increase the half-life of this enzyme and thus stimulate an increase in its activity in vivo. It also protects ornithine decarboxylase against proteolysis by chymotrypsin in vitro.

Kinetics of Various 99mTc-Sn-Pyrophosphate Compounds in the Rat

1977 ◽  
Vol 16 (04) ◽  
pp. 157-162 ◽  
Author(s):  
C. Schümichen ◽  
B. Mackenbrock ◽  
G. Hoffmann
Keyword(s):  
Normal Saline ◽  
Half Life ◽  
Compound A ◽  
Short Half Life ◽  
Low Concentrations ◽  
The Stability ◽  
Kinetics Of ◽  
In Vitro Stability

SummaryThe bone-seeking 99mTc-Sn-pyrophosphate compound (compound A) was diluted both in vitro and in vivo and proved to be unstable both in vitro and in vivo. However, stability was much better in vivo than in vitro and thus the in vitro stability of compound A after dilution in various mediums could be followed up by a consecutive evaluation of the in vivo distribution in the rat. After dilution in neutral normal saline compound A is metastable and after a short half-life it is transformed into the other 99mTc-Sn-pyrophosphate compound A is metastable and after a short half-life in bone but in the kidneys. After dilution in normal saline of low pH and in buffering solutions the stability of compound A is increased. In human plasma compound A is relatively stable but not in plasma water. When compound B is formed in a buffering solution, uptake in the kidneys and excretion in urine is lowered and blood concentration increased.It is assumed that the association of protons to compound A will increase its stability at low concentrations while that to compound B will lead to a strong protein bond in plasma. It is concluded that compound A will not be stable in vivo because of a lack of stability in the extravascular space, and that the protein bond in plasma will be a measure of its in vivo stability.

scholarly journals Mechanism for Stabilizing mRNAs Involved in Methanol-Dependent Methanogenesis of Cold-Adaptive Methanosarcina mazei zm-15

2013 ◽  
Vol 80 (4) ◽  
pp. 1291-1298 ◽  
Author(s):  
Yi Cao ◽  
Jie Li ◽  
Na Jiang ◽  
Xiuzhu Dong
Keyword(s):  
Half Life ◽  
Content Type ◽  
Methanosarcina Mazei ◽  
Reverse Transcription Quantitative Pcr ◽  
Aceticlastic Methanogenesis ◽  
The Stability ◽  
Zoige Wetland ◽  
Mrna Half Life

ABSTRACTMethylotrophic methanogenesis predominates at low temperatures in the cold Zoige wetland in Tibet. To elucidate the basis of cold-adapted methanogenesis in these habitats,Methanosarcina mazeizm-15 was isolated, and the molecular basis of its cold activity was studied. For this strain, aceticlastic methanogenesis was reduced 7.7-fold during growth at 15°C versus 30°C. Methanol-derived methanogenesis decreased only 3-fold under the same conditions, suggesting that it is more cold adaptive. Reverse transcription-quantitative PCR (RT-qPCR) detected <2-fold difference in the transcript abundances ofmtaA1,mtaB1, andmtaC1, the methanol methyltransferase (Mta) genes, in 30°C versus 15°C culture, whileackAandptamRNAs, encoding acetate kinase (Ack) and phosphotransacetylase (Pta) in aceticlastic methanogenesis, were 4.5- and 6.8-fold higher in 30°C culture than in 15°C culture. Thein vivohalf-lives ofmtaA1andmtaC1B1mRNAs were similar in 30°C and 15°C cultures. However, thepta-ackAmRNA half-life was significantly reduced in 15°C culture compared to 30°C culture. Using circularized RNA RT-PCR, large 5′ untranslated regions (UTRs) (270 nucleotides [nt] and 238 nt) were identified formtaA1andmtaC1B1mRNAs, while only a 27-nt 5′ UTR was present in thepta-ackAtranscript. Removal of the 5′ UTRs significantly reduced thein vitrohalf-lives ofmtaA1andmtaC1B1mRNAs. Remarkably, fusion of themtaA1ormtaC1B15′ UTRs topta-ackAmRNA increased itsin vitrohalf-life at both 30°C and 15°C. These results demonstrate that the large 5′ UTRs significantly enhance the stability of the mRNAs involved in methanol-derived methanogenesis in the cold-adaptiveM. mazeizm-15.

scholarly journals Stabilization of the mRNA for the uncoupling protein thermogenin by transcriptional/translational blockade and by noradrenaline in brown adipocytes differentiated in culture: a degradation factor induced by cessation of stimulation?

1994 ◽  
Vol 302 (1) ◽  
pp. 81-86 ◽  
Author(s):  
C Picó ◽  
D Herron ◽  
A Palou ◽  
A Jacobsson ◽  
B Cannon ◽  
...  
Keyword(s):  
Half Life ◽  
Uncoupling Protein ◽  
Mrna Level ◽  
Direct Interaction ◽  
Brown Adipocytes ◽  
Degradation Factor ◽  
The Stability ◽  
High Level

The stability of the mRNA coding for the uncoupling protein thermogenin was investigated in mouse brown-fat cells differentiated in culture. After 7 days in culture, the cells were stimulated for 24 h with noradrenaline, and a high level of thermogenin mRNA was then observed. If noradrenaline treatment was continued, the mRNA level remained high, but, upon withdrawal of noradrenaline, the level decreased rapidly, with a half-life of only 2.7 h. The presence of transcriptional (actinomycin) or translational (cycloheximide) inhibitors prolonged the apparent half-life by about 50%. The presence of noradrenaline during transcriptional blockade led to a further stabilization of thermogenin mRNA. It was concluded that an induced (or short-lived) gene product is important for thermogenin mRNA degradation. Direct interaction of noradrenaline with the cultured brown adipocytes could apparently not mimic the paradoxical destabilization of thermogenin mRNA in vivo, previously observed in the cold-exposed mouse [Jacobsson, Cannon and Nedergaard (1987) FEBS Lett. 244, 353-356], indicating significant differences between the systems in vitro and in vivo.

scholarly journals DL-α-Monofluoromethylputrescine is a potent irreversible inhibitor of Escherichia coli ornithine decarboxylase

1982 ◽  
Vol 204 (3) ◽  
pp. 771-775 ◽  
Author(s):  
A Kallio ◽  
P P McCann ◽  
P Bey
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Ornithine Decarboxylase ◽  
Half Life ◽  
Arginine Decarboxylase ◽  
Biosynthetic Enzyme ◽  
Irreversible Inhibitor ◽  
E Coli

DL-alpha-Monofluoromethylputrescine (compound R.M.I. 71864) is an enzyme-activated irreversible inhibitor of the biosynthetic enzyme ornithine decarboxylase from Escherichia coli. This compound, however, has much less effect in vitro on ornithine decarboxylase obtained from Pseudomonas aeruginosa. These findings are in contrast with those previously found with the substrate analogue DL-alpha-difluoromethylornithine (compound R.M.I. 71782). The K1 of the DL-alpha-monofluoromethylputrescine for the E. coli ornithine decarboxylase is 110 microM, and the half-life (t1/2) calculated for an infinite concentration of inhibitor is 2.1 min. When DL-alpha-monofluoromethylputrescine is used in combination with DL-alpha-difluoromethylarginine (R.M.I. 71897), an irreversible inhibitor of arginine decarboxylase, in vivo in E. coli, both decarboxylase activities are inhibited (greater than 95%) but putrescine levels are only decreased to about one-third of control values and spermidine levels are slightly increased.

scholarly journals Designing and Development of a Tandem Bivalent Nanobody against VEGF165

2021 ◽  
Author(s):  
Farnaz Khodabakhsh ◽  
Morteza Salimian ◽  
Pardis Ziaee ◽  
Fatemeh Kazemi-Lomedasht ◽  
Mahdi Behdani ◽  
...  
Keyword(s):  
Antiproliferative Activity ◽  
Half Life ◽  
Pharmacokinetic Parameters ◽  
Native Protein ◽  
Metal Affinity ◽  
Significant Difference ◽  
Inhibition Of Angiogenesis ◽  
The Stability

Background: Inhibition of angiogenesis using monoclonal antibodies is an effective strategy in cancer therapy. However, they could not penetrate sufficiently into solid tumors. Antibody fragments have solved this issue. However, they suffer from short in vivo half-life. In the current study, a tandem bivalent strategy was used to enhance the pharmacokinetic parameters of an anti-VEGF165 nanobody. Methods: Homology modeling and MD simulation were used to check the stability of protein. The cDNA was cloned into pHEN6C vector and the expression was investigated in WK6 Escherichia coli (E. coli) cells by SDS-PAGE and western blot. After purification, the size distribution of tandem bivalent nanobody was investigated by dynamic light scattering. Moreover, in vitro antiproliferative activity and pharmacokinetic study were studied in HUVECs and Balb/c mice, respectively. Results: RMSD analysis revealed the tandem bivalent nanobody had good structural stability after 50 ns of simulation. A hinge region of llama IgG2 was used to fuse the domains. The expression was induced by 1 mM IPTG at 25°C for overnight. A 30 kDa band in 12% polyacrylamide gel and nitrocellulose paper has confirmed the expression. The protein was successfully purified using metal affinity chromatography. MTT assay revealed there is no significant difference between the antiproliferative activity of tandem bivalent nanobody and the native protein. The hydrodynamic radius and terminal half-life of tandem bivalent nanobody increased approximately 2-fold by multivalency compared to the native protein. Conclusion: Our data revealed that the physicochemical as well as in vivo pharmacokinetic parameters of tandem bivalent nanobody was significantly improved.

Formation and Structure of Casein Micelles in Lactating Mammary Tissue

1970 ◽  
Vol 28 ◽  
pp. 150-151
Author(s):  
Robert J. Carroll ◽  
Marvin P. Thompson ◽  
Harold M. Farrell
Keyword(s):  
Size Distribution ◽  
G Protein ◽  
Milk Proteins ◽  
Mammary Tissue ◽  
Colloidal System ◽  
Casein Micelles ◽  
The Stability

Milk is an unusually stable colloidal system; the stability of this system is due primarily to the formation of micelles by the major milk proteins, the caseins. Numerous models for the structure of casein micelles have been proposed; these models have been formulated on the basis of in vitro studies. Synthetic casein micelles (i.e., those formed by mixing the purified αsl- and k-caseins with Ca2+ in appropriate ratios) are dissimilar to those from freshly-drawn milks in (i) size distribution, (ii) ratio of Ca/P, and (iii) solvation (g. water/g. protein). Evidently, in vivo organization of the caseins into the micellar form occurs in-a manner which is not identical to the in vitro mode of formation.

Half-Life of Platelet Factor 4 (PF-4) in Plasma and Platelets from Macaca Mulatta

1977 ◽  
Vol 37 (01) ◽  
pp. 073-080 ◽  
Author(s):  
Knut Gjesdal ◽  
Duncan S. Pepper
Keyword(s):  
Macaca Mulatta ◽  
Half Life ◽  
Human Platelet ◽  
Gel Filtration ◽  
Platelet Factor 4 ◽  
Active Protein ◽  
Platelet Factor ◽  
Membrane Bound

SummaryHuman platelet factor 4 (PF-4) showed a reaction of complete identity with PF-4 from Macaca mulatta when tested against rabbit anti-human-PF-4. Such immunoglobulin was used for quantitative precipitation of in vivo labelled PF-4 in monkey serum. The results suggest that the active protein had an intra-platelet half-life of about 21 hours. In vitro 125I-labelled human PF-4 was injected intravenously into two monkeys and isolated by immuno-precipita-tion from platelet-poor plasma and from platelets disrupted after gel-filtration. Plasma PF-4 was found to have a half-life of 7 to 11 hours. Some of the labelled PF-4 was associated with platelets and this fraction had a rapid initial disappearance rate and a subsequent half-life close to that of plasma PF-4. The results are compatible with the hypothesis that granular PF-4 belongs to a separate compartment, whereas membrane-bound PF-4 and plasma PF-4 may interchange.

Applicability of Instability Index for In vitro Protein Stability Prediction

2019 ◽  
Vol 26 (5) ◽  
pp. 339-347 ◽  
Author(s):  
Dilani G. Gamage ◽  
Ajith Gunaratne ◽  
Gopal R. Periyannan ◽  
Timothy G. Russell
Keyword(s):  
Protein Stability ◽  
Caulobacter Crescentus ◽  
Effect Of Temperature ◽  
Instability Index ◽  
Dipeptide Composition ◽  
Experimental Conditions ◽  
The Stability ◽  
Vitro Protein

Background: The dipeptide composition-based Instability Index (II) is one of the protein primary structure-dependent methods available for in vivo protein stability predictions. As per this method, proteins with II value below 40 are stable proteins. Intracellular protein stability principles guided the original development of the II method. However, the use of the II method for in vitro protein stability predictions raises questions about the validity of applying the II method under experimental conditions that are different from the in vivo setting. Objective: The aim of this study is to experimentally test the validity of the use of II as an in vitro protein stability predictor. Methods: A representative protein CCM (CCM - Caulobacter crescentus metalloprotein) that rapidly degrades under in vitro conditions was used to probe the dipeptide sequence-dependent degradation properties of CCM by generating CCM mutants to represent stable and unstable II values. A comparative degradation analysis was carried out under in vitro conditions using wildtype CCM, CCM mutants and two other candidate proteins: metallo-β-lactamase L1 and α -S1- casein representing stable, borderline stable/unstable, and unstable proteins as per the II predictions. The effect of temperature and a protein stabilizing agent on CCM degradation was also tested. Results: Data support the dipeptide composition-dependent protein stability/instability in wt-CCM and mutants as predicted by the II method under in vitro conditions. However, the II failed to accurately represent the stability of other tested proteins. Data indicate the influence of protein environmental factors on the autoproteolysis of proteins. Conclusion: Broader application of the II method for the prediction of protein stability under in vitro conditions is questionable as the stability of the protein may be dependent not only on the intrinsic nature of the protein but also on the conditions of the protein milieu.

scholarly journals Screening marine algae metabolites as high-affinity inhibitors of SARS-CoV-2 main protease (3CLpro): an in silico analysis to identify novel drug candidates to combat COVID-19 pandemic

2020 ◽  
Vol 63 (1) ◽  
Author(s):  
Ghazala Muteeb ◽  
Adil Alshoaibi ◽  
Mohammad Aatif ◽  
Md. Tabish Rehman ◽  
M. Zuhaib Qayyum
Keyword(s):  
Hydrophobic Interactions ◽  
In Silico Analysis ◽  
Salt Bridge ◽  
Competitive Inhibitor ◽  
Binding Energies ◽  
Dynamics Simulation ◽  
In Vivo Studies ◽  
Energy Calculation

AbstractThe recent dissemination of SARS-CoV-2 from Wuhan city to all over the world has created a pandemic. COVID-19 has cost many human lives and created an enormous economic burden. Although many drugs/vaccines are in different stages of clinical trials, still none is clinically available. We have screened a marine seaweed database (1110 compounds) against 3CLpro of SARS-CoV-2 using computational approaches. High throughput virtual screening was performed on compounds, and 86 of them with docking score <  − 5.000 kcal mol−1 were subjected to standard-precision docking. Based on binding energies (< − 6.000 kcal mol−1), 9 compounds were further shortlisted and subjected to extra-precision docking. Free energy calculation by Prime-MM/GBSA suggested RC002, GA004, and GA006 as the most potent inhibitors of 3CLpro. An analysis of ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) properties of RC002, GA004, and GA006 indicated that only RC002 (callophysin A, from red alga Callophycus oppositifolius) passed Lipinski’s, Veber’s, PAINS and Brenk’s filters and displayed drug-like and lead-like properties. Analysis of 3CLpro-callophysin A complex revealed the involvement of salt bridge, hydrogen bonds, and hydrophobic interactions. callophysin A interacted with the catalytic residues (His41 and Cys145) of 3CLpro; hence it may act as a mechanism-based competitive inhibitor. Docking energy and docking affinity of callophysin A towards 3CLpro was − 8.776 kcal mol−1 and 2.73 × 106 M−1, respectively. Molecular dynamics simulation confirmed the stability of the 3CLpro-callophysin A complex. The findings of this study may serve as the basis for further validation by in vitro and in vivo studies.

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