Interaction of [125I]? nerve growth factor with acidic proteins

Rusenko, Kirt W.; Stach, Robert W.

doi:10.1007/bf00964044

Cited by 20 publications

(11 citation statements)

References 25 publications

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“…Internalization of EGF, unlike insulin, is unaffected by the transglutaminase inhibitors methylamine and dansylcadaverine [Pastan and Willingham, 19811, perhaps because a majority of EGF receptors already exist in an "aggregated" (clustered) state prior to exposure of the cells to EGF [Haigler et al, 1979;Linsley and Fox, 1980bl. It may be that the lack of an effect of transglutaminase inhibitors on sequestration, covalent NGF-NGFR complex formation and internalization is due to the NGFR-1's already being in an aggregated or clustered state [Rusenko, 1982;Stach and Rusenko, 19841. The NGF-NGFR covalent complex is held together by disulfide bonds since dithiothreitol (DTT) reduction abolishes the complex [Stach and Rusenko, 1984;Compito et al, 19861. Pre-incubation of cells with sulfhydryl reagents, such as N-ethylamaleimide, iodoacetamide, or 5,5'-dithiobis (2-nitrobenzoic acid) not only prevents formation of the covalent NGF-NGFR complex, but also inhibits the biological response to NGF [Compito et al, 19861.…”

Section: Covalent Attachment Of Ngf To Its Receptormentioning

confidence: 99%

Binding of nerve growth factor to its receptor

Stach

Perez‐Polo

1987

J of Neuroscience Research

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Section: Covalent Attachment Of Ngf To Its Receptormentioning

confidence: 99%

Binding of nerve growth factor to its receptor

Stach

Perez‐Polo

1987

J of Neuroscience Research

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“…7 s NGF was isolated by a modification of the method of Stach et al [ 19771 as described by Rusenko and Stach [1981]. PNGF was isolated from 7s NGF as described by Stach et a1 [1979] and was stored at 4°C in 0.2% acetic acid at a concentration of 2-4 mg/ml.…”

Section: Isolation Of Ngfmentioning

confidence: 99%

Covalent attachment of ¹²⁵I‐β nerve growth factor to its receptors on sympathetic neurons

Stach

Rusenko

1984

J of Neuroscience Research

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When 125I-beta nerve growth factor binds to sympathetic and sensory neurons, some labeled ligand is sequestered (becomes inaccessible to the external milieu) in a time- and energy-dependent manner. It would appear that the higher affinity receptor (type I) participates in this process to a greater extent than does the lower affinity receptor (type II) [ Olender and Stach , 1980; Olender et al., 1981]. A small portion of the sequestered 125I-beta nerve growth factor is found as part of a high molecular weight complex. When cells, which have been incubated with 125I-beta nerve growth factor, are solubilized with Triton X-100 and subjected to sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis, a complex with an apparent molecular weight of approximately 240,000 is obtained. The formation of the covalent complex can be prevented by the prior addition of excess unlabeled beta nerve growth factor or sodium fluoride and dinitrophenol. The covalent 125I-beta nerve growth factor-receptor complex is dissociated in 50 mM dithiothreitol indicating that disulfide linkages are involved. At concentrations of beta nerve growth factor (3.8 X 10(-11) -3.8 X 10(-10) M) where maximal fiber outgrowth occurs in vitro, approximately 50-266 attomoles (0.3-1.6% of the type I receptors) of the covalent complex are formed per 10(7) nerve cells. These data suggest that a small portion of the 125I-beta nerve growth sequestered by sympathetic neurons becomes covalently attached to its receptor subsequent to its sequestration in a manner which appears to involve type I receptors.

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“…The question of whether the low-affinity binding (NGFR 11) or the high-affinity binding (NGFR I) is relevant to N G F action is unresolved. There is evidence that high-affinity NGFR is necessary for neurite elongation (Calissano and Shelanski, 1980;Carbonetto and Stach, 1982;Olender and Stach, 1980;Rusenko and Stach, 1981;Hosang and Shooter, 1985;Ishii, 1982, 1985). There is also evidence that the two binding activities may be related in PC 12 (Landreth and Shooter, 1980).…”

mentioning

confidence: 99%

Nerve Growth Factor Receptors in Human Neuroblastoma Cells

Marchetti

Perez‐Polo

1987

Journal of Neurochemistry

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Receptors for the nerve growth factor protein (NGFR) present in the human neuroblastoma cell line LAN-1 were characterized. LAN-1 cells display high-affinity (type I, with KD value of 5.9 X 10(-11) M) and low-affinity (type II, with KD value of 9.2 X 10(-9) M) binding to NGF. NGFR were fractionated by preparative isoelectric focusing in a granulated gel (PEGG). High-affinity binding was found in the 5.9-6.2 pH region of the PEGG, and low-affinity binding in the 4.6-4.8 and 8.8-9.3 pH ranges. After further analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) we observed both 92.5- and 200-kDa molecular species associated with NGF binding activity. The 200-kDa protein was found in fractions displaying high-affinity NGF binding and the 92.5-kDa protein in fractions displaying low-affinity NGF binding. Equilibrium binding analysis of NGF in PEGG fractions confirmed the presence of two specific saturable binding sites with KD values similar to those observed for whole dissociated cells. When NGFR II activity from the acidic region of the PEGG chromatogram was incubated with NGFR II from the basic region of the PEGG chromatogram, there was no change in NGF binding or in the number of apparent NGF receptors. However, incubation of these same fractions with a fraction having only NGFR I showed an apparent increase in high-affinity NGF binding and a decrease in low-affinity NGF binding. Immunoprecipitation of this "mixed" fraction and analysis on SDS-PAGE under reduced and nonreduced conditions showed 200-kDa and 92.5-kDa proteins under nonreduced conditions and a 92.5-kDa protein under reduced conditions. Our findings are consistent with the hypothesis that there are two distinct NGF receptors in NGF-responsive cells. The interconvertibility of low- and high-affinity receptors and the possible existence of a modulator type protein or of "silent" type receptors are also in agreement with our findings.

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Interaction of [125I]? nerve growth factor with acidic proteins

Cited by 20 publications

References 25 publications

Binding of nerve growth factor to its receptor

Binding of nerve growth factor to its receptor

Covalent attachment of ¹²⁵I‐β nerve growth factor to its receptors on sympathetic neurons

Nerve Growth Factor Receptors in Human Neuroblastoma Cells

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Interaction of [125I]? nerve growth factor with acidic proteins

Cited by 20 publications

References 25 publications

Binding of nerve growth factor to its receptor

Binding of nerve growth factor to its receptor

Covalent attachment of 125I‐β nerve growth factor to its receptors on sympathetic neurons

Nerve Growth Factor Receptors in Human Neuroblastoma Cells

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Covalent attachment of ¹²⁵I‐β nerve growth factor to its receptors on sympathetic neurons