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1.
Navolotskaia EV Zargarova TA Lepikhova TN Turobov VL Nurieva RI Malkova NV Zav'ialov VP Lipkin VM 《Bioorganicheskaia khimiia》2000,26(1):31-38
The antiproliferative and immunosuppressive in vitro effects of immunocortin, a synthetic adrenocorticotropin-like (ACTH-like) decapeptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH, whose sequence corresponds to segment 11-20 of the variable part of the human IgG1 heavy chain, were studied. At concentrations of 10(-11)-10(-7) M, immunocortin was found to inhibit the growth of the human MT-4 T-lymphoblastoid cell line, to suppress the blast transformation of thymocytes, and to decrease the spontaneous mobility of peritoneal macrophages and their bactericidal action toward the virulent strain Salmonella typhimurium 415. By using a 125I-labeled "addressing" fragment of ACTH ?[125I]ACTH-(13-24)?, we showed that MT-4 cells express specific receptors for ACTH (Kd 97 pM). Immunocortin and human ACTH (but not the heavy chain of IgG1) competitively inhibited the binding of [125I]ACTH-(13-24) to these receptors with Ki1 of 0.38 and Ki2 of 0.34 nM, respectively. Specific receptors for ACTH (Kd 5.8 nM) on mouse thymocytes were detected and characterized. The unlabeled immunocortin was shown to complete with labeled ACTH-(13-24) for binding to these receptors (Ki = 1.8 nM) and this binding of immunocortin to receptors on thymocytes activates adenylate cyclase from these cells and increases the intracellular concentration of cAMP. 相似文献
2.
E. V. Navolotskaya V. I. Vanina T. A. Zargarova E. N. Goncharenko N. Yu. Kudryashova M. Ya. Akhalaya V. B. Sadovnikov S. G. Semushina A. A. Kolobov E. A. Kampe-Nemm V. V. Yurovskii V. M. Lipkin 《Russian Journal of Bioorganic Chemistry》2004,30(4):315-319
The effect of immunocortin, an ACTH-like decapeptide VKKPGSSVKV corresponding to the 11–20 sequence of the variable part of the human IgG1 heavy chain on the content of 11-hydroxycorticosteroids (CS) in rat adrenal glands and blood serum in vivo was studied. An intramuscular injection of immunocortin at a dose of 10 g/kg was found in an hour to induce a twofold decrease in CS content in the adrenal glands and a 1.8-fold increase in the blood serum CS content. At the same time, an immunocortin dose of 100 g/kg exerted practically no effect on the CS content and its dose of 1000 g/kg increased the CS content both in adrenal glands and in blood serum by 1.6 and 2.2 times, respectively. Four hours after the injection of any of the three doses of immunocortin, the CS content in adrenal glands did not differ from the control value, and after 24 h the content decreased threefold. Immunocortin was shown to be bound by the ACTH receptors in the membranes of the rat adrenal cortex with a high affinity and specificity (inhibiting the specific binding of 125I-labeled ACTH-(11–24) peptide with K
i of 1.2 nM). 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.
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
5.
Guanine nucleotides modulate the function of chemotactic cyclic AMP receptors inDictyostelium discoideum 总被引:3,自引:0,他引:3
Pim M. W. Janssens Peter L. J. van der Geer Jos C. Arents Roel van Driel 《Molecular and cellular biochemistry》1985,67(2):119-124
Summary Guanosine di- and triphosphates specifically decrease the affinity of chemotactic cAMP receptors in isolatedDictyostelium discoideum membranes. The K0.5 was increased from 50 nM to 150 nM. Receptors were shown to be heterogeneous in dissociation kinetics. In the absence of
guanine nucleotides three dissociation processes could be resolved, having first order rate constants of 8.7 x 10−4, 1.3 X 10−2, and higher than 0.1 s−1. Guanine nucleotides decreased the affinity for cAMP by transforming the slowest dissociating receptor form (KD is 8 nM) to forms dissociating more rapidly. Our data indicate that a guanine nucleotide binding protein (G-protein) is involved
in the transduction of the cAMP signal inD. discoideum. 相似文献
6.
Lepikhova TN Navolotskaya EV Zargarova TA Nurieva RI Lipkin VM Zav'yalov VP 《Peptides》2000,21(3):353-357
Influence of the ACTH-like peptide H-Val-Lys-Lys-Pro-Gly-Ser-Ser-Val-Lys-Val-OH corresponding to the sequence 11-20 of the variable part of human immunoglobulin G1 (IgG1) heavy chain on growth of MT-4 human T-lymphoblastoid cell line was investigated. It was found that the ACTH-like peptide at concentration range 10(-11) -10(-7) M inhibits the proliferation of MT-4 cells. Labeled ACTH 'address segment' [(125)I]ACTH (13-24) was used to establish that MT-4 cells express specific receptors for ACTH (K(d) = 97 pM). The ACTH-like peptide and human ACTH (but not IgG1 heavy chain) were shown to compete with [(125)I]ACTH (13-24) for binding to these receptors (K(i1) = 0.38 nM and K(i2) = 0.34 nM). 相似文献
7.
The tachykinins, substance P (SP) and neurokinin A (NKA), are agonists for the NK1 and NK2 receptors, respectively. Tachykinins have various respiratory effects, including bronchoconstriction. This study characterizes
tachykinin binding sites in the rabbit lung. We hypothesize that (2-[125I]iodohistidyl1)Neurokinin A ([125I]NKA) interacts with NK1 and NK2 binding sites in the rabbit lung. The Kd determined from saturation isotherms was 0.69 X/÷1.14 nM (geometic mean X/÷ SEM) and the Bmax was 4.15±0.22 femtomole/mg protein (arithmetic mean±SEM). Competitive inhibition studies with NKA, SP and various selective
tachykinin agonists showed the rank order of potency: [β-Ala8]-Neurokinin A 4–10=SP ≫ NKA ≫ [Sar9,Met(O2)11]-Substance P. [β-Ala8]-Neurokinin A 4–10, a selective NK2 agonist, and SP inhibition of [125I]NKA binding were best described using a two-site model. Competitive inhibition studies using the selective nonpeptide NK2 antagonist (SR 48968) and the selective nonpeptide NK1 antagonist (CP 96,345) revealed Ki's of 5.5 nM and 8.1 nM, respectively. Our data therefore suggest that [125I]NKA binds to both the NK1 and NK2 receptors in the lung.
Special issue dedicated to Dr. Kinya Kuriyama. 相似文献
8.
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
9.
Wada T Imanishi T Kawaguchi A Mori MX Mori Y Imoto K Ichida S 《Neurochemical research》2005,30(8):1045-1054
Characteristics for the specific binding of 125I-ω-CTX GVIA and 125I-ω-CTX MVIIC to crude membranes from BHKN101 cells expressing the α1B subunits of Cav2.2 channels and from mice brain lacking the α1B subunits of Cav2.2 channels, particularly, the effects of CaM and various Ca2+ channel blockers on these specific bindings were investigated. Specific binding of 125I-ω-CTX GVIA to the crude membranes from BHKN101 cells was observed, but not from control BHK6 cells. ω-CTX GVIA, ω-CTX MVIIC
and ω-CTX SVIB inhibited the specific binding of 125I-ω-CTX GVIA to crude membranes from BHKN101 cells, and the IC50 values for ω-CTXGVIA, ω-CTX MVIIC and ω-CTX SVIB were 0.07, 8.5 and 1.7 nM, respectively. However, ω-agatoxin IVA and calciseptine
at concentrations of 10−9–10−6 M did not inhibit specific binding. Specific binding was also about 80% inhibited by 20 μg protein/ml CaM. The amount of
125I-ω-CTX GVIA (30 pM) specifically bound to membranes from brain of knockout mice lacking α1B subunits of Cav2.2 channels was about 30% of that to the crude membranes from brain of wild-type. On the other hand, specific binding of
125I-ω-CTX MVIIC (200 pM) was observed on the crude membranes of both BHKN101 and control BHK6 cells. The specific binding of
125I-ω-CTX MVIIC (200 pM) was not inhibited by ω-CTX GVIA and ω-CTX SVIB, and also ω-Aga IVA and calciseptine at concentrations
of 10−9–10−7 M, although specific binding was almost completely dose dependently inhibited by non-radiolabeled ω-CTX MVIIC (IC50 value was about 0.1 nM). 20 μg protein/ml CaM did not inhibit specific binding. Therefore, these results suggest that BHKN101
cells have a typical Cav2.2 channels which are also inhibited by CaM and have not specific binding sites for ω-CTX MVIIC, although ω-CTX MVIIC is
a blocker for both Cav2.1 (α1A; P/Q-type) and Cav2.2 channels. 相似文献
10.
Navolotskaya E. V. Malkova N. V. Lepikhova T. N. Krasnova S. B. Zargarova T. A. Zav'yalov V. P. Lipkin V. M. 《Russian Journal of Bioorganic Chemistry》2001,27(5):318-322
The synthetic peptide SLTCLVKGFY, corresponding to the 364–373 amino acid sequence of the human IgG heavy chain (Immunorphin), was found to compete with [125I] -endorphin for binding by high-affinity receptors on T lymphocytes isolated from the blood of healthy donors (K
i0.6 nM). The fragments 3–10, 4–10, 5–10, and 6–10 of Immunorphin also inhibited the binding (K
i2.2, 3.4, 8.0, and 15 nM, respectively). Specificity of these receptors was studied: they turned out to be insensitive to naloxone and [Met]enkephaline and, therefore, are not opioid. The K
dvalues of the specific binding of 125I-labeled Immunorphin and its 6–10 fragment to the receptor were found to be 7.4 and 36.3 nM, respectively. 相似文献
11.
Isabel A. Lüthy Enrique T. Segura Viviana I. Lüthy Eduardo H. Charreau Ricardo S. Calandra 《Journal of comparative physiology. B, Biochemical, systemic, and environmental physiology》1985,155(5):611-614
Summary (125I)-ovine prolactin (oPRL) binding was found in several brain areas of the toad,Bufo arenarum Hensel. The olfactory bulb, cerebral hemispheres, and both dorsal and ventral mesencephalic regions showed saturable, high
affinity, (125I)-oPRL binding, ranging between 5.6 to 29.9 fmol/mg protein, while the association constant (K
a) by Scatchard analysis was between 4.0 to 8.7×109 M−1. This binding was compared with the Scatchard plot of the kidney, which has been already described by other groups, and gave
41.7 fmol/mg protein andK
a 2.5×109 M−1. Liver showed no binding and in the cerebral hemispheres (125I)-oPRL was not displaced by non-lactogenic hormones, indicating that binding was hormone and tissue specific. 相似文献
12.
We investigated the block of KATP channels by glibenclamide in inside-out membrane patches of rat flexor digitorum brevis muscle.
(1) We found that glibenclamide inhibited KATP channels with an apparent K
i
of 63 nm and a Hill coefficient of 0.85. The inhibition of KATP channels by glibenclamide was unaffected by internal Mg2+.
(2) Glibenclamide altered all kinetic parameters measured; mean open time and burst length were reduced, whereas mean closed
time was increased.
(3) By making the assumption that binding of glibenclamide to the sulphonylurea receptor (SUR) leads to channel closure, we
have used the relation between mean open time, glibenclamide concentration and K
D
to estimate binding and unbinding rate constants. We found an apparent rate constant for glibenclamide binding of 9.9 × 107
m
−1 sec−1 and an unbinding rate of 6.26 sec−1.
(4) Glibenclamide is a lipophilic molecule and is likely to act on sulfonylurea receptors from within the hydrophobic phase
of the cell membrane. The glibenclamide concentration within this phase will be greater than that in the aqueous solution
and we have taken this into account to estimate a true binding rate constant of 1.66 × 106
m
−1 sec−1.
Received: 7 July 1996/Revised: 4 October 1996 相似文献
13.
Navolotskaia EV Vanina VI Zargarova TA Goncharenko EN Kudriashova NIu Akhalaia MIa Sadovnikov VB Semushkina SG Kolobov AA Kampe-Nemm EA Iurovskiĭ VV Lipkin VM 《Bioorganicheskaia khimiia》2004,30(4):350-355
The effect of immunocortin, an ACTH-like decapeptide VKKPGSSVKV corresponding to the 11-20 sequence of the variable part of the human IgG1 heavy chain on the content of 11-hydroxycorticosteroids (CS) in rat adrenal glands and blood serum in vivo was studied. An intramuscular injection of immunocortin at a dose of 10 microg/kg was found in an hour to induce a twofold decrease in CS content in the adrenal glands and a 1.8-fold increase in the blood serum CS content. At the same time, an immunocortin dose of 100 microg/kg exerted practically no effect on the CS content and its dose of 1000 microg/kg increased the CS content both in adrenal glands and in blood serum by 1.6 and 2.2 times, respectively. Four hours after the injection of any of the three doses of immunocortin, the CS content in adrenal glands did not differ from the control value, and after 24 h the content decreased threefold. Immunocortin was shown to be bound by the ACTH receptors in the membranes of the rat adrenal cortex with a high affinity and specificity (inhibiting the specific binding of 125I-labeled ACTH-(11-24) peptide with Ki of 1.2 nM). 相似文献
14.
Willem Bintig Judith Baumgart Wilhelm J. Walter Alexander Heisterkamp Holger Lubatschowski Anaclet Ngezahayo 《Journal of bioenergetics and biomembranes》2009,41(1):85-94
Purinergic signalling in rat GFSHR-17 granulosa cells was characterised by Ca2+-imaging and perforated patch-clamp. We observed a resting intracellular Ca2+-concentration ([Ca2+]i) of 100 nM and a membrane potential of −40 mV. This was consistent with high K+− and Cl− permeability and a high intracellular Cl− concentration of 40 mM. Application of ATP for 5–15 s every 3 min induced repeated [Ca2+]i increases and a 30 mV hyperpolarization. The phospholipase C inhibitor U73122 or the IP3-receptor antagonist 2-aminoethoethyl diphenyl borate suppressed ATP responses. Further biochemical and pharmacological experiments
revealed that ATP responses were related to stimulation of P2Y2 and P2Y4 receptors and that the [Ca2+]i increase was a prerequisite for hyperpolarization. Inhibitors of Ca2+-activated channels or K+ channels did not affect the ATP-evoked responses. Conversely, inhibitors of Cl− channels hyperpolarized cells to −70 mV and suppressed further ATP-evoked hyperpolarization. We propose that P2Y2 and P2Y4 receptors in granulosa cells modulate Cl− permeability by regulating Ca2+-release. 相似文献
15.
Kayvan Ariani Mark W. Hamblin Grace L. Tan Carol A. Stratford Dr. Roland D. Ciaranello 《Neurochemical research》1989,14(9):835-843
Several manipulations that affect G protein/receptor coupling also alter the binding of [125I]iodocyanopindolol ([125I]ICYP)±cyanopindolol (±CYP) to rat brain 5-HT1B binding sites in radiologand binding assays. Inclusion of 5 mM MgSO4 in these assays results in a small but significant increase in the affinity of [125I]ICYP (fromK
D=0.046 nM toK
D=0.037 nM). In contrast, 100 M Gpp(NH)p, GTP, or GDP reduce [125I]ICYP affinity (K
D=0.056 nM with GTP) while ATP and GMP are less effective.±CYP affinity for 5-HT1B sites labeled by [3H]dihydroergotamine ([3H]DE) also displays a small but significant reduction (from Ki=1.4 nM to Ki=3.5nM) by the inclusion of 100 M GTP. Pre-treatment of the brain membranes with N-ethylmaleimide (NEM) in concentrations known to inactivate many G proteins reduces 5-HT1B specific binding of [125I]ICYP. The NEM induced reduction in [125I]ICYP binding can be reversed by reconstitution with purified exogenous G proteins (Go and Gi), demonstrating directly that high affinity binding of [125I]ICYP to 5-HT1B sites is dependent on G proteins. The effects of Mg2+ ion, guanine nucleotides, NEM and G protein reconstitution on [125I]ICYP and ±CYP binding are all hallmarks of agonist binding to G protein linked receptors. The effect of GTP, however, is quantitatively much less for the binding of these pindolol derivatives than for the binding of 5-HT, a presumed full agonist at 5-HT1B sites. The relatively slight stabilization of [125I]ICYP and ±CYP binding conferred by G protein/5-HT1B receptor interaction may reflect the molecular events underlying previous observations that these compounds are partial 5-HT1B agoinists. 相似文献
16.
Single-channel properties of a delayed rectifier voltage-gated K+ channel (I-type) were investigated in peripheral myelinated axons from Xenopus laevis. Channels activated between −60 and −40 mV with a potential of half-maximal activation, E50, at −47.5 mV. Averaged single-channel currents activated with a time delay at all membrane potentials tested. Time to half-maximal
activation decreased from 80 to 1.6 msec between −60 and +40 mV. The channel inactivated monoexponentially with a time constant
of 10.9 sec at −40 mV. The time constant of deactivation was 126 msec at −80 mV and 16.9 msec at −110 mV. In symmetrical 105
mm K+, the single-channel conductance (γ) was 22 and 13 pS at negative and positive membrane potentials, respectively, at 13–15°C.
In Na+-rich solution with 2.5 mm extracellular K+γ was 7 pS and the reversal potential was negative to −80 mV, indicating a high selectivity for K+ over Na+. γ depended on extracellular K+ concentration (K
D
= 19.6 mm) and temperature (Q
10= 1.45). External tetraethylammonium (TEA) reduced the apparent single-channel current amplitude at all potentials tested
with a half-maximal inhibiting concentration (IC50) of 0.6 mm. Open probability of the channel, but not single-channel current amplitude was decreased by extracellular dendrotoxin (DTX,
IC50= 6.8 nm) and mast cell degranulating peptide (MCDP, IC50= 41.9 nm). In Ringer solution the membrane potential of macroscopic I-channel patches was about −65 mV and depolarized under TEA and
DTX. It is concluded that besides their activation during action potentials, I-channels may also stabilize the resting membrane
potential.
Received: 2 June 1995/Revised: 13 October 1995 相似文献
17.
Kiyoshi Nokihara Yoshihiro Nakata Tadashi Yasuhara Victor Wray 《International journal of peptide research and therapeutics》2008,14(1):52-57
Although there is no amino acid sequence similarity between maxadilan (Maxa) and pituitary adenylate cyclase activating polypeptide
(PACAP), our synthetic Maxa was found to bind PACAP specific receptors (PAC-1 receptors) with a high affinity, but low potency
for the accumulation of cAMP in PC12 cells. Competitive binding studies of 125I-PACAP-27 to rat cortical membranes allowed exploration of the structural requirements for this interaction using mini-libraries
constructed by solid-phase peptided synthesis, that include disulfide isomers, N-, C- and middle segment deleted peptides
and analogs. Maxa as well as PACAP38 inhibited the specific binding of 125I-PACAP-27 with IC50 values of 3.89 and 4.90 nM, respectively. The most potent derivative of our synthetic Maxa-analogs with an IC50 value of 1.99 nM was Maxa[1–23 + 43–61, S–S14–51 Ala1,5] which consists of N- (position 1–23) and C- (position 43-61) terminal linear fragments cross-linked by a disulfide bridge
between positions 14 and 51. This peptide did not increase intracellular cAMP, at a concentration of 100 nM, but inhibited
cAMP accumulation induced by 1 nM PACAP-27 in PC12 cells, whereas wild Maxa increased intracellular cAMP although it was weaker
than PACAP-27. Our data suggest deletion of the middle segment between residues 24–42 affords some derivatives that behave
as low affinity antagonists.
This paper is dedicated to the memory of Professor Bruce Merrifield, a pioneer and one of the most important contributors
to solid-phase synthesis. 相似文献
18.
Ian A. Darby Jacob Bouhnik Ericque D. Coezy Pierre Corvol 《In vitro cellular & developmental biology. Animal》1991,27(1):21-24
Summary Binding characteristics and effects of 3,5,-3′-triiodo-l-thyronine (T3) on angiotensinogen production in HepG2 were studied in serum-free medium. Binding was performed on intact cells and on partially purified isolated nuclei using
[125I]T3. Scatchard plots revealed one class of high affinity binding sites with a Kd of approximately 80 pmol/liter. Calculation of maximum binding showed that HepG2 possess approximately 1000 binding sites per cell. Unlabeled T3 and T4 competed for binding sites on intact HepG2 with 50% inhibition of [125I]T3 binding at approximately 3.0 and 38.0 pmol/liter, respectively. The HepG2 showed a dose-dependent increase in angiotensinogen production in serum-free medium which was maximal at 10−5 mol/liter (two-fold increase/106 cells/24 h) and had an EC50 of approximately 5.0×10−8 mol/liter. T3 also produced after 24 h a dose-dependent increase in DNA highly correlated with T3 applied (r=0.88,P<0.01). In conclusion, this study shows that HepG2 possess specific high affinity binding sites for T3 and that T3 stimulates angiotensinogen production and DNA synthesis in these cells.
Dr. Darby is supported by INSERM (France)/NH and MRC (Australia) exchange fellowship. 相似文献
19.
P. O. Vardevanyan A. P. Antonyan G. A. Manukyan A. T. Karapetyan A. K. Shchyolkina O. F. Borisova 《Molecular Biology》2000,34(2):272-276
Isotherms of the EtBr adsorption on native and denatured poly(dA)poly(dT) in the temperature interval 20–70°C were obtained.
The EtBr binding constants and the number of binding sites were determined. The thermodynamic parameters of the EtBr intercalation
complex upon changes of solution temperature 20–48°C were calculated: 1.0·106 M−1≤K≤1.4·106 M−1, free energy ΔG
o=−8.7±0.3 kcal/mol, enthalpy ΔH
o≅0, and entropy ΔS
o=28±0.5 cal/(mol deg). UV melting has shown that the melting temperature (T
m) of EtBr-poly(dA)poly(dT) complexes (μ=0.022,4.16·10−5 M EtBr) increased by 17°C as compared with the ΔT
m of free homopolymer, whereas the half-width of the transition (T
m) is not changed. It was shown for the first time that EtBr forms complexes of two types on single-stranded regions of poly(dA)poly(dT)
denatured at 70°C: strong (K
1=1.7·105 M−1; ΔG
o=−8.10±0.03 kcal/mol) and weak (K
2=2.9·103 M−1; ΔG
o=−6.0±0.3 kcal/mol).The ΔG
o of the strong and weak complexes was independent of the solution ionic strength, 0.0022≤μ≤0.022. A model of EtBr binding
with single-stranded regions of poly(dA)poly(dT) is discussed. 相似文献
20.
L. A. Nesterova B. N. Manukhin 《Biochemistry (Moscow) Supplemental Series A: Membrane and Cell Biology》2011,5(1):85-91
The influence of isoprenaline- and propranolole-induced activation and inhibition of β-adrenoreceptors on the specific nonselective
α2-antagonist [3H]RX821002 binding was studied on rat cerebral cortex subcellular membrane fractions. It was shown that the ligand-receptor
interaction for α2-adrenoreceptors corresponded to the model that assumed the presence of one receptor pool and binding of two ligand molecules
to a receptor dimer. The following parameters were determined for [3H]RX821002 binding to α2-adrenoreceptors: K
d1 = 1.57 ± 0.27 nM, B
max = 7.24 ± 1.63 fmol/mg of protein, n = 2. In the case of isoprenaline-induced activation of β-adrenoreceptors the binding of radiolabeled ligand to α2-adrenoreceptors was described by the same model. The affinity of α2-adrenoreceptors for [3H]RX821002 decreased more than twofold (K
d = 3.55 ± 0.02 nM) and the quantity of active receptors increased by 69% (B
max = 12.24 ± 0.06 fmol/mg of protein). Propranolole changed the model of ligand binding, and two pools of receptors were detected
with the following parameters: K
d1 = 0.61 ± 0.02 nM, K
d2 = 3.41 ± 0.13 nM, B
ml = 1.88 ± 0.028 fmol/mg of protein, B
m2 = 9.27 ± 0.08 fmol/mg of protein, n = 2. The data suggest that α2-adrenoreceptors in subcellular membrane fractions from rat cerebral cortex exist in dimeric form. Isoprenaline and propranolole
exhibit modulating effect on the specific antagonist binding to α2-adrenoreceptors, which results in the inhibition and alteration of [3H]RX821002 binding parameters. 相似文献