共查询到20条相似文献,搜索用时 78 毫秒
1.
Lipase-catalyzed synthesis of isoamyl acetate in hexane at 10–250 MPa at 80°C and 1–100 MPa at 40°C resulted in activation
volumes of −12.9 ± 1.7 and −21.6 ± 2.9 cm3 mol−1, respectively. Increasing pressure from 10 to 200 MPa resulted in approximately 10-fold increase in V
max at both 40 and 80°C. Pressure increased the K
m from 2.4 ± 0.004 to 38 ± 0.78 mM at 40°C. In contrast, at 80°C the pressure did not affect the K
m. 相似文献
2.
Ramaiah Sivakumar Giriyapura R. Vijayakumar Balaraman Manohar 《Biocatalysis and Biotransformation》2013,31(4):299-305
Kinetic studies of two glucosylation reactions catalyzed by an amyloglucosidase from Rhizopus sp. leading to the synthesis of vanillin-α/β-D-glucoside from D-glucose and vanillin and curcumin-bis-α-D-glucoside from D-glucose and curcumin were investigated in detail. Initial reaction rates were determined from kinetic runs involving different concentrations of D-glucose and vanillin (5?mM to 0.1?M) or D-glucose and curcumin (5?mM to 0.1?M). Graphical double reciprocal plots showed that the kinetics of the two enzyme catalyzed reactions exhibited Ping-Pong Bi-Bi mechanism where competitive substrate inhibition by vanillin/curcumin led to dead-end amyloglucosidase–vanillin/curcumin complexes at higher concentrations of vanillin/curcumin. An attempt to obtain the best fit of this kinetic model through computer simulation yielded in good approximation, the values of four important kinetic parameters, vanillin-α/β-D-glucoside: kcat=35.0±3.2 10?5M?h?1·mg, Ki=10.5±1.1?mM, KmD-glucose=60.0±6.2?mM, Kmvanillin=50.0±4.8?mM; curcumin-bis-α-D-glucoside: kcat=6.07±0.58 10?5M?h?1·mg, Ki=3.0±0.28?mM, KmD-glucose=10.0±0.9?mM, Kmcurcumin=4.6±0.5?mM. 相似文献
3.
Elena V. Navolotskaya Yulia A. Kovalitskaya Vladimir B. Sadovnikov Yury A. Zolotarev Alexander A. Kolobov Vladimir V. Yurovsky Valery M. Lipkin 《International journal of peptide research and therapeutics》2008,14(1):10-15
Synthetic adrenocorticotropic hormone (ACTH)-like octapeptide leukocorticotropin (GKVLKKRR), corresponding to the amino acid
sequence 81–88 of pro-interleukin-1α, was labeled with tritium (specific activity of 22 Ci/mmol) and was found to bind to
rat adrenal cortex membranes with high affinity and specificity (K
d = 2.2 ± 0.1 nM). Synthetic 125I-labeled ACTH fragment 11–24 was also obtained (specific activity of 98 Ci/mmol) and shown to bind to ACTH receptor on rat
adrenal cortex with high affinity (K
d = 1.8 ± 0.1 nM). Unlabeled leukocorticotropin was found to actively replace 125I-labeled ACTH (11–24) in the receptor-ligand complex (K
i = 2.0 ± 0.1 nM). Leukocorticotropin at concentration range of 1–1000 nМ did not affect the adenylate cyclase activity in
adrenocortical membranes. Thus, leukocorticotropin is an antagonist of ACTH receptor.
ACTH-like peptide GKVLKKRR is an antagonist of ACTH
This work financeed by the Russian Foundation for Basic Research (grant No. 05-04-48060), by the programs Leading Scientific
Schools (grant No. 312.2003.4), Molecular and Cellular Biology (chairman V.M. Lipkin), and Naukogrady (grant No. 04-04-97200),
and by the International Science and Technology Center (project No. 2615). V.V. Yurovsky is supported by the American Heart
Association (grant No. 0555415U). 相似文献
4.
Alexander A. Kolobov Nikolai I. Kolodkin Cynthia Tuthill Yury A. Zolotarev Elena V. Navolotskaya 《International journal of peptide research and therapeutics》2008,14(2):97-103
Tritium-labeled dipeptide bestim (γ-D-Glu-L-Trp) with a specific activity of 45 Ci/mmol was obtained by the high-temperature solid-state catalytic isotope exchange (HSCIE)
reaction. [3H]bestim was found to bind with high affinity to mouse peritoneal macrophages (K
d
2.1 ± 0.1 nM) and thymocytes (K
d
3.1 ± 0.2 nM) and also plasma membranes isolated from these cells (K
d
18.6 ± 0.2 and 16.7 ± 0.3 nM respectively). The specific bonding of [3H]bestim with macrophages and thymocytes was inhibited by unlabeled dipeptide thymogen (L-Glu-L-Trp) (K
i
0.9 ± 0.1 and 1.1 ± 0.1 nM respectively). Treatment of the macrophages and thymocytes with trypsin led to their loss of capacity
to bind [3H]bestim. Bestim at concentrations range of 0.1–1000 nМ reduced the adenylate cyclase activity in macrophage and thymocyte
membranes. 相似文献
5.
Yuliia N. Nekrasova Vladimir B. Sadovnikov Yury A. Zolotarev Elena V. Navolotskaya 《Journal of peptide science》2010,16(6):263-268
The synthetic peptide TPLVTLFK corresponding to the sequence 12–19 of β‐endorphin (referred to as octarphin) was found to bind to high‐affinity naloxone‐insensitive binding sites on membranes isolated from the rat brain cortex (Kd = 2.6 ± 0.2 nM ). The binding specificity study revealed that these binding sites were insensitive not only to naloxone but also to α‐endorphin, γ‐endorphin, [Met5]enkephalin, and [Leu5]enkephalin, as well. The [3H]octarphin specific binding with brain membranes was inhibited by unlabeled β‐endorphin (Ki = 2.4 ± 0.2 nM ) and a selective agonist of nonopioid β‐endorphin receptor decapeptide immunorphin SLTCLVKGFY (Ki = 2.9 ± 0.2 nM ). At the same time, unlabeled octarphin completely (by 100%) inhibited the specific binding of [3H]immunorphin with membranes (Ki = 2.8 ± 0.2 nM ). Thus, octarphin binds with a high affinity and specificity to nonopioid receptor of β‐endorphin on rat brain cortex membranes. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
6.
Yu. A. Kovalitskaya A. A. Kolobov E. A. Kampe-Nemm Yu. A. Zolotarev V. V. Yurovskii V. B. Sadovnikov V. M. Lipkin E. V. Navolotskaya 《Russian Journal of Bioorganic Chemistry》2008,34(1):24-29
Tritium-labeled synthetic fragments of human adrenocorticotropic hormone (ACTH) [3H]ACTH (11–24) and [3H]ACTH (15–18) with a specific activity of 22 and 26 Ci/mmol, respectively, were obtained. It was found that [3H]ACTH-(11–24) binds to membranes of the rat adrenal cortex with high affinity and high specificity (K d 1.8 ± 0.1 nM). Twenty nine fragments of ACTH (11–24) were synthesized, and their ability to inhibit the specific binding of [3H]ACTH (11–24) to adrenocortical membranes was investigated. The shortest active peptide was found to be an ACTH fragment (15–18) (KKRR) (K i 2.3 ± 0.2 nM), whose [3H] labeled derivative binds to rat adrenocortical membranes (K d 2.1 ± 0.1 nM) with a high affinity. The specific binding of [3H]ACTH-(15–18) was inhibited by 100% by unlabeled ACTH (11–24) (K i 2.0 ± 0.1 nM). ACTH (15–18) in the concentration range of 1–1000 nM did not affect the adenylate cyclase activity of adrenocortical membranes and, therefore, is an antagonist of the ACTH receptor. 相似文献
7.
Elena V. Navolotskaya Yulia A. Kovalitskaya Yury A. Zolotarev Vladimir B. Sadovnikov 《Journal of peptide science》2008,14(10):1121-1128
Selective agonist of nonopioid β‐endorphin receptor decapeptide immunorphin (SLTCLVKGFY) was labeled with tritium (the specific activity of 24 Ci/mmol). [3H]Immunorphin was found to bind to nonopioid β‐endorphin receptor of mouse peritoneal macrophages (Kd = 2.0 ± 0.1 nM ). The [3H]immunorphin specific binding with macrophages was inhibited by unlabeled β‐endorphin (Ki = 2.9 ± 0.2 nM ) and was not inhibited by unlabeled naloxone, α‐endorphin, γ‐endorphin and [Met5]enkephalin (Ki > 10 µM ). Thirty fragments of β‐endorphin have been synthesized and their ability to inhibit the [3H]immunorphin specific binding to macrophages was studied. Unlabeled fragment 12–19 (TPLVTLFK, the author's name of the peptide octarphin) was found to be the shortest peptide possessing practically the same inhibitory activity as β‐endorphin (Ki = 3.1 ± 0.3 nM ). The peptide octarphin was labeled with tritium (the specific activity of 28 Ci/mmol). [3H]Octarphin was found to bind to macrophages with high affinity (Kd = 2.3 ± 0.2 nM ). The specific binding of [3H]octarphin was inhibited by unlabeled immunorphin and β‐endorphin (Ki = 2.4 ± 0.2 and 2.7 ± 0.2 nM , respectively). Copyright © 2008 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
8.
A. A. Kolobov N. I. Kolodkin Yu. A. Zolotarev C. Tuthill E. V. Navolotskaya 《Russian Journal of Bioorganic Chemistry》2008,34(1):37-42
The tritium-labeled dipeptide bestim (γ-D-Glu-L-Trp) with a specific activity of 45 Ci/mmol was obtained by high-temperature solid-state catalytic isotope exchange. It was found that [3H]bestim binds with a high affinity to murine peritoneal macrophages (K d 2.1 ± 0.1 nM) and thymocytes (K d 3.1 ± 0.2 nM), as well as with plasma membranes isolated from these cells (K d 18.6 ± 0.2 and 16.7 ± 0.3 nM, respectively). The specific binding of [3H]bestim to macrophages and thymocytes was inhibited by the unlabeled dipeptide thymogen (L-Glu-L-Trp) (K i 0.9 ± 0.1 and 1.1 ± 0.1 nM, respectively). After treatment with trypsin, macrophages and thymocytes lost the ability to bind [3H]bestim. Bestim in the concentration range of 10?10 to 10?6 M reduced the adenylate cyclase activity in the membranes of murine macrophages and thymocytes. 相似文献
9.
Uur Tutar Ümit M. Koyiit Hayreddin Gezegen 《Journal of biochemical and molecular toxicology》2019,33(4)
A series of 1,3‐bis‐chalcone derivatives ( 3a‐i, 6a‐i and 8 ) were synthesized and evaluated antimicrobial, antibiofilm and carbonic anhydrase inhibition activities. In this evaluation, 6f was found to be the most active compound showing the same effect as the positive control against Bacillus subtilis and Streptococcus pyogenes in terms of antimicrobial activity. Biofilm structures formed by microorganisms were damaged by compounds at the minimum inhibitory concentration value between 0.5% and 97%.1,3‐bis‐chalcones ( 3a‐i, 6a‐i and 8 ) showed good inhibitory action against human (h) carbonic anhydrase (CA) isoforms I and II. hCA I and II were effectively inhibited by these compounds, with K i values in the range of 94.33 ± 13.26 to 787.38 ± 82.64 nM for hCA I, and of 100.37 ± 11.41 to 801.76 ± 91.11 nM for hCA II, respectively. In contrast, acetazolamide clinically used as CA inhibitor showed K i value of 1054.38 ± 207.33 nM against hCA I, and 983.78 ± 251.08 nM against hCA II, respectively. 相似文献
10.
In this work, the inhibitory activity of Voriconazole was measured against some metabolic enzymes, including human carbonic anhydrase (hCA) I and II isoenzymes, acetylcholinesterase (AChE), and α‐glycosidase; the results were compared with standard compounds including acetazolamide, tacrine, and acarbose. Half maximal inhibition concentration (IC50) values were obtained from the enzyme activity (%)‐[Voriconazole] graphs, whereas Ki values were calculated from the Lineweaver‐Burk graphs. According to the results, the IC50 value of Voriconazole was 40.77 nM for α‐glycosidase, while the mean inhibition constant (Ki) value was 17.47 ± 1.51 nM for α‐glycosidase. The results make an important contribution to drug design and have pharmacological applications. In addition, the Voriconazole compound demonstrated excellent inhibitory effects against AChE and hCA isoforms I and II. Voriconazole had Ki values of 29.13 ± 3.57 nM against hCA I, 15.92 ± 1.90 nM against hCA II, and 10.50 ± 2.46 nM against AChE. 相似文献
11.
A selective agonist of non-opioid β-endorphin receptor synthetic peptide octarphin (TPLVTLFK, specific activity 28 Ci/mmol)
was prepared. The [3H]octarphin binding to rat myocardium membranes before and after experimental myocardial infarction (EMI) was studied. It
was found that [3H]octarphin with high affinity and specificity binds to non-opioid β-endorphin receptor of rat myocardium membranes before
EMI: K
d1 value of the [3H]octarphin specific binding to membranes was 1.8 ± 0.2 nM. In 3 h after EMI a sharp lowering in affinity of the binding is
observed (K
d2 = 13.3 ± 0.4 nM), and in 48 h its almost complete restoration (K
d4 = 2.2 ± 0.3 nM). The results indicate participation of non-opioid β-endorphin receptor in the regulation of myocardial activity. 相似文献
12.
Smooth muscle cells isolated from the secondary and tertiary branches of the rabbit mesenteric artery contain large Ca2+-dependent channels. In excised patches with symmetrical (140 mm) K+ solutions, these channels had an average slope conductance of 235 ± 3 pS, and reversed in direction at −6.1 ± 0.4 mV. The
channel showed K+ selectivity and its open probability (P
o
) was voltage-dependent. Iberiotoxin (50 nm) reversibly decreased P
o
, whereas tetraethylammonium (TEA, at 1 mm) reduced the unitary current amplitude. Apamin (200 nm) had no effect. The channel displayed sublevels around 1/3 and 1/2 of the mainstate level. The effect of [Ca2+] on P
o
was studied and data fitted to Boltzmann relationships. In 0.1, 0.3, 1.0 and 10 μm Ca2+, V
1/2 was 77.1 ± 5.3 (n= 18), 71.2 ± 4.8 (n= 16), 47.3 ± 10.1 (n= 11) and −14.9 ± 10.1 mV (n= 6), respectively. Values of k obtained in 1 and 10 μm [Ca2+] were significantly larger than that observed in 0.1 μm [Ca2+]. With 30 μm NS 1619 (a BKCa channel activator), V
1/2 values were shifted by 39 mV to the left (hyperpolarizing direction) and k values were not affected. TEA applied intracellularly, reduced the unitary current amplitude with a K
d
of 59 mm. In summary, BKCa channels show a particularly weak sensitivity to intracellular TEA and they also display large variation in V
1/2 and k. These findings suggest the possibility that different types (isoforms) of BKCa channels may exist in this vascular tissue.
Received: 22 December 1997/Revised: 27 March 1998 相似文献
13.
Feifan Du Max A. Kruziki Elizabeth J. Zudock Yi Zhang Patrick S. Lown Benjamin J. Hackel 《Biotechnology and bioengineering》2019,116(3):526-535
The Gp2 domain is a 45 amino-acid scaffold that has been evolved for specific, high-affinity binding towards multiple targets and was proven useful in molecular imaging and biological antagonism. It was hypothesized that Gp2 may benefit from increased hydrophilicity for improved physiological distribution as well as for physicochemical robustness. We identified seven exposed hydrophobic sites for hydrophilic mutations and experimentally evaluated single mutants, which yielded six mutations that do not substantially hinder expression, binding affinity or specificity (to epidermal growth factor receptor), and thermal stability. Eight combinations of these mutations improved hydrophilicity relative to the parental Gp2 clone as assessed by reverse-phase high-performance liquid chromatography (p < 0.05). Secondary structures and refolding abilities of the selected single mutants and all multimutants were unchanged relative to the parental ligand. A variant with five hydrophobic-to-hydrophilic mutations was identified with enhanced solubility as well as reasonable binding affinity ( K d = 53–63 nM), recombinant yield (1.3 ± 0.8 mg/L), and thermal stability ( T m = 53 ± 3°C). An alternative variant with a cluster of three leucine-to-hydrophilic mutations was identified with increased solubility, nominally increased binding affinity ( K d = 13–28 nM) and reasonable thermal stability ( T m = 54.0 ± 0.6°C) but reduced yield (0.4 ± 0.3 mg/L). In addition, a ≥7°C increase in the midpoint of thermal denaturation was observed in one of the single mutants (T21N). These mutants highlight the physicochemical tradeoffs associated with hydrophobic-to-hydrophilic mutation within a small protein, improve the solubility and hydrophilicity of an existent molecular imaging probe, and provide a more hydrophilic starting point for discovery of new Gp2 ligands towards additional targets. 相似文献
14.
Synthesis and investigation of the conversion reactions of pyrimidine‐thiones with nucleophilic reagent and evaluation of their acetylcholinesterase,carbonic anhydrase inhibition,and antioxidant activities
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Parham Taslimi Afsun Sujayev Fikret Turkan Emin Garibov Zübeyir Huyut Vagif Farzaliyev Sevgi Mamedova İlhami Gulçin 《Journal of biochemical and molecular toxicology》2018,32(2)
The conversion reactions of pyrimidine‐thiones with nucleophilic reagent were studied during this scientific research. For this purpose, new compounds were synthesized by the interaction between 1,2‐epoxy propane, 1,2‐epoxy butane, and 4‐chlor‐1‐butanol and pyrimidine‐thiones. These pyrimidine‐thiones derivatives ( A–K ) showed good inhibitory action against acetylcholinesterase (AChE), and human carbonic anhydrase (hCA) isoforms I and II. AChE inhibition was in the range of 93.1 ± 33.7–467.5 ± 126.9 nM. The hCA I and II were effectively inhibited by these compounds, with Ki values in the range of 4.3 ± 1.1–9.1 ± 2.7 nM for hCA I and 4.2 ± 1.1–14.1 ± 4.4 nM for hCA II. On the other hand, acetazolamide clinically used as CA inhibitor showed Ki value of 13.9 ± 5.1 nM against hCA I and 18.1 ± 8.5 nM against hCA II. The antioxidant activity of the pyrimidine‐thiones derivatives ( A–K ) was investigated by using different in vitro antioxidant assays, including Cu2+ and Fe3+ reducing, 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH?) radical scavenging, and Fe2+ chelating activities. 相似文献
15.
The kinetics of NADH oxidation by the outer membrane electron transport system of intact beetroot (Beta vulgaris L.) mitochondria were investigated. Very different values for Vmax and the Km for NADH were obtained when either antimycin A-insensitive NADH-cytochrome c activity (Vmax= 31 ± 2.5 nmol cytochrome c (mg protein)?1 min?1; Km= 3.1 ± 0.8 μM) or antimycin A-insensitive NADH-ferricyanide activity (Vmax= 1.7 ± 0.7 μmol ferricyanide (mg protein)?1 min?1; Km= 83 ± 20 μM) were measured. As ferricyanide is believed to accept electrons closer to the NADH binding site than cytochrome c, it was concluded that 83 ± 20 μM NADH represented a more accurate estimate of the binding affinity of the outer membrane dehydrogenase for NADH. The low Km determined with NADH-cytochrome c activity may be due to a limitation in electron flow through the components of the outer membrane electron transport chain. The Km for NADH of the externally-facing inner membrane NADH dehydrogenase of pea leaf (Pisum sativum L. cv. Massey Gem) mitochondria was 26.7 ± 4.3 μM when oxygen was the electron acceptor. At an NADH concentration at which the inner membrane dehydrogenase should predominate, the Ca2+ chelator, ethyleneglycol-(β-aminoethylether)-N,N,-tetraacetic acid (EGTA), inhibited the oxidation of NADH through to oxygen and to the ubiquinone-10 analogues, duroquinone and ubiquinone-1, but had no effect on the antimycin A-insensitive ferricyanide reduction. It is concluded that the site of action of Ca2+ involves the interaction of the enzyme with ubiquinone and not with NADH. 相似文献
16.
The Ca2+-activated maxi K+ channel was found in the apical membrane of everted rabbit connecting tubule (CNT) with a patch-clamp technique. The mean
number of open channels (NP
o
) was markedly increased from 0.007 ± 0.004 to 0.189 ± 0.039 (n= 7) by stretching the patch membrane in a cell-attached configuration. This activation was suggested to be coupled with the
stretch-activation of Ca2+-permeable cation channels, because the maxi K+ channel was not stretch-activated in both the cell-attached configuration using Ca2+-free pipette and in the inside-out one in the presence of 10 mm EGTA in the cytoplasmic side. The maxi K+ channel was completely blocked by extracellular 1 μm charybdotoxin (CTX), but was not by cytoplasmic 33 μm arachidonic acid (AA). On the other hand, the low-conductance K+ channel, which was also found in the same membrane, was completely inhibited by 11 μm AA, but not by 1 μm CTX. The apical K+ conductance in the CNT was estimated by the deflection of transepithelial voltage (ΔV
t
) when luminal K+ concentration was increased from 5 to 15 mEq. When the tubule was perfused with hydraulic pressure of 0.5 KPa, the ΔV
t
was only −0.7 ± 0.4 mV. However, an increase in luminal fluid flow by increasing perfusion pressure to 1.5 KPa markedly enhanced
ΔV
t
to −9.4 ± 0.9 mV. Luminal application of 1 μm CTX reduced the ΔV
t
to −1.3 ± 0.6 mV significantly in 6 tubules, whereas no significant change of ΔV
t
was recorded by applying 33 μm AA into the lumen of 5 tubules (ΔV
t
=−7.2 ± 0.5 mV in control vs.ΔV
t
=−6.7 ± 0.6 mV in AA). These results suggest that the Ca2+-activated maxi K+ channel is responsible for flow-dependent K+ secretion by coupling with the stretch-activated Ca2+-permeable cation channel in the rabbit CNT.
Received: 21 August 1997/Revised: 20 March 1998 相似文献
17.
Akira Katarao Takahiro Mune Katsutada Takahashi 《Bioscience, biotechnology, and biochemistry》2013,77(9):2443-2449
The effects of streptomycin, tetracycline and chloramphenicol on the growth of Escherichia coli were studied quantitatively in a conduction-type batch calorimeter at pH 6.2 and 37°C. Change in the growth thermograms with increasing drug concentrations in the medium were analyzed with a kinetic model of non-competitive inhibition.The number of drug molecules needed to inactivate a unit viable cell, m, was estimated to be 1.2 + 0.1, 0.7 ±0.1 and 1.3 ±0.1 for streptomycin, tetracycline and chloramphenicol, respectively. With the assumption that the drug binding sites are all identical, the microscopic dissociation constant per binding site, K, for the drugs was found to be 0.19, 0.43 and 0.94μmoldm-3 for streptomycin, tetracycline and chloramphenicol, respectively. With these m and K{ values, drug potency curves were drawn for the three drugs. 相似文献
18.
Anne H. ODonnell Xiaojie Yao Larry D. Byers 《Biochimica et Biophysica Acta - Proteins and Proteomics》2004,1703(1):63-67
The solvent kinetic isotope effects (SKIE) on the yeast α-glucosidase-catalyzed hydrolysis of p-nitrophenyl and methyl-d-glucopyranoside were measured at 25 °C. With p-nitrophenyl-d-glucopyranoside (pNPG), the dependence of kcat/Km on pH (pD) revealed an unusually large (for glycohydrolases) solvent isotope effect on the pL-independent second-order rate constant, DOD(kcat/Km), of 1.9 (±0.3). The two pKas characterizing the pH profile were increased in D2O. The shift in pKa2 of 0.6 units is typical of acids of comparable acidity (pKa=6.5), but the increase in pKa1 (=5.7) of 0.1 unit in going from H2O to D2O is unusually small. The initial velocities show substrate inhibition (Kis/Km~200) with a small solvent isotope effect on the inhibition constant [DODKis=1.1 (±0.2)]. The solvent equilibrium isotope effects on the Kis for the competitive inhibitors d-glucose and α-methyl d-glucoside are somewhat higher [DODKi=1.5 (±0.1)]. Methyl glucoside is much less reactive than pNPG, with kcat 230 times lower and kcat/Km 5×104 times lower. The solvent isotope effect on kcat for this substrate [=1.11 (±0. 02)] is lower than that for pNPG [=1.67 (±0.07)], consistent with more extensive proton transfer in the transition state for the deglucosylation step than for the glucosylation step. 相似文献
19.
Harinder S. Hundal Peter M. Taylor Nicholas M. Willhoft Bryan Mackenzie Sylvia Y. Low Martyn R. Ward Michael J. Rennie 《生物化学与生物物理学报:疾病的分子基础》1992,1180(2):137-146
We wished to examine the effects of diabetes on muscle glutamine kinetics. Accordingly, female Wistar rats (200 g) were made diabetic by a single injection of streptozotocin (85 mg/kg) and studied 4 days later; control rats received saline. In diabetic rats, glutamine concentration of gastrocnemius muscle was 33% less than in control rats: 2.60 ± 0.06 μmol/g vs. 3.84 ± 0.13 μmol/g (P < 0.001). In gastrocnemius muscle, glutamine synthetase activity (Vmax) was unaltered by diabetes (approx. 235 nmol/min per g) but glutaminase Vmax increased from 146 ± 29 to 401 ± 94 nmol/min per g; substrate Km values of neither enzyme were affected by diabetes. Net glutamine efflux (AZ concentration difference × blood flow) from hindlimbs of diabetic rats in vivo was greater than control values (?30.0 ± 3.2 vs. ?1.9 ± 2.6 nmol/min per g (P < 0.001) and hindlimb NH3 uptake was concomitantly greater (about 27 nmol/min per g). The glutamine transport capacity (Vmax) of the Na-dependent System Nm in perfused hindlimb muscle was 29% lower in diabetic rats than in controls (820 ± 50 vs. 1160 ± 80 nmol/min per g (P < 0.01)), but transporter Km was the same in both groups (9.2 ± 0.5 nM). The difference between inward and net glutamine fluxes indicated that glutamine efflux in perfused hindlimbs was stimulated in diabetes at physiological perfusate glutamine (0.5 mM); ammonia (1 mM in perfusate) had little effect on net glutamine flux in control and diabetic muscles. In Intramuscular Na+ was 26% greater in diabetic (13.2 μmol/g) than control muscle, but muscle K+ (100 μmol/g) was similar. The accelerated rate of glutamine release from skeletal muscle and the lower muscle free glutamine concentration observed in diabetes may result from a combination of; (i), a diminished Na+ electrochemical gradient (i.e., the net driving force for glutamine accrual in muscle falls); (ii), a faster turnover of glutamine in muscle and (iii), an increased Vmax/Km for sarcolemmal glutamine efflux. 相似文献
20.
We have investigated the interaction of two peptides (ShB — net charge +3 and ShB:E12KD13K — net charge +7) derived from the NH2-terminal domain of the Shaker K+ channel with purified, ryanodine-modified, cardiac Ca2+-release channels (RyR). Both peptides produced well resolved blocking events from the cytosolic face of the channel. At a
holding potential of +60 mV the relationship between the probability of block and peptide concentration was described by a
single-site binding scheme with 50% saturation occurring at 5.92 ± 1.06 μm for ShB and 0.59 ± 0.14 nm for ShB:E12KD13K. The association rates of both peptides varied with concentration (4.0 ± 0.4 sec−1μm
−1 for ShB and 2000 ± 200 sec−1μm
−1 for ShB:E12KD13K); dissociation rates were independent of concentration. The interaction of both peptides was influenced by applied
potential with the bulk of the voltage-dependence residing in Koff. The effectiveness of the inactivation peptides as blockers of RyR is enhanced by an increase in net positive charge. As
is the case with inactivation and block of K+ channels, this is mediated by a large increase in Kon. These observations are consistent with the proposal that the conduction pathway of RyR contains negatively charged sites
which will contribute to the ion handling properties of this channel.
Received: 15 December 1997/Revised: 13 March 1998 相似文献