Chymotrypsin substrate analogues inhibit endocytosis of insulin and insulin receptors in adipocytes.

Jochen, Albert; Berhanu, Paulos

doi:10.1083/jcb.103.5.1807

Cited by 17 publications

(11 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Membranebound metalloproteases have been found in animal cells, which play important roles in intracellular vesicular transport including endocytosis (Jochen and Berhanu, 1986;Strous et al, 1988) and exocytosis (Mundy and Strittmatter, 1985) as well as the transport between endoplasmic reticulum and Golgi complex (Strous et al, 1988). Membranebound metalloproteases have been found in animal cells, which play important roles in intracellular vesicular transport including endocytosis (Jochen and Berhanu, 1986;Strous et al, 1988) and exocytosis (Mundy and Strittmatter, 1985) as well as the transport between endoplasmic reticulum and Golgi complex (Strous et al, 1988).…”

Section: Discussionmentioning

confidence: 99%

“…Recent studies have implicated some metalloproteases in a number of important intracellular processes. Membranebound metalloproteases have been found in animal cells, which play important roles in intracellular vesicular transport including endocytosis (Jochen and Berhanu, 1986;Strous et al, 1988) and exocytosis (Mundy and Strittmatter, 1985) as well as the transport between endoplasmic reticulum and Golgi complex (Strous et al, 1988). Mitochondrial processing protease is a soluble metalloprotease, and participates in the transport of protein precursors into the organelles.…”

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Purification and characterization of a processing protease from rat liver mitochondria.

et al. 1989

View full text Add to dashboard Cite

A processing protease has been purified from the matrix fraction of rat liver mitochondria. The purified protease contained two protein subunits of 55 kd (P‐55) and 52 kd (P‐52) as determined by SDS‐PAGE. The processing protease was estimated to be 105 kd in gel filtration, indicating that the two protein subunits form a heterodimeric complex. At high ionic conditions, the two subunits dissociated. The purified processing protease cleaved several mitochondrial protein precursors destined to different mitochondrial compartments, including adrenodoxin, malate dehydrogenase, P‐450(SCC) and P‐450(11 beta), but the processing efficiencies were different each other. The endoprotease nature of the processing protease was confirmed with the purified enzyme using adrenodoxin precursor as the substrate; both the mature form and the extension peptide were detected after the processing. The processing activity of the protease was inhibited by metal chelators, and reactivated by Mn2+, indicating that the protease is a metalloprotease.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Purification and characterization of a processing protease from rat liver mitochondria.

et al. 1989

View full text Add to dashboard Cite

show abstract

“…10) and 2 mM-Gly-Phe-amide (results not shown) which do not inhibit metalloendoproteinases. The metalloendoproteinase inhibitors 1,10-phenanthroline (0.5 mM) and phosphoramidon (20 pUM), the calpain inhibitors antipain and leupeptin (1 mm each [37]) and the chymotrypsin inhibitors phenylmethanesulphonyl fluoride (0.1 mM), N-acetyl-L-Tyr ethyl ester (1 mM, [38]) and aprotinin (up to 1 unit/ml) were without any effect on Ca21_ stimulated ouabain binding (results not shown). .…”

Section: Proteinase Inhibitorsmentioning

confidence: 99%

Micromolar free calcium exposes ouabain-binding sites in digitonin-permeabilized Xenopus laevis oocytes

Schmalzing

Kröner

1990

Biochemical Journal

View full text Add to dashboard Cite

As demonstrated previously, digitonin-permeabilized Xenopus oocytes have a large internal pool of sodium pumps which are inaccessible to cytosolic ouabain [Schmalzing, Kröner & Passow (1989) Biochem. J. 260, 395-399]. Access to internal ouabain-binding sites required permeabilization of inner membranes with SDS. In the present study, micromolar free Ca2+ was found to stimulate ouabain binding in the digitonin-permeabilized cells (K0.5 0.5 microM-Ca2+, h 1.9, average of seven experiments) without disrupting intracellular membranes. Sustained incubation at 9 microM-Ca2+ was as effective as SDS in inducing access to the ouabain-binding sites of the internal sodium pumps. Omission of either Mg2+ or ATP completely abolished the Ca2+ effect. Half-maximal stimulation by Ca2+ required approx. 0.4 mM-MgATP. Of a variety of nucleotides tested, none was as effective as ATP (rank order ATP greater than ADP greater than ATP[S] (adenosine 5'-[gamma-thio]triphosphate) greater than CTP greater than UTP greater than ITP = XTP greater than GTP). Pi, AMP, cyclic AMP, cyclic GMP, GTP[S] (guanosine 5'-[gamma-thio]triphosphate) and a stable ATP analogue p[NH]ppA (adenosine 5'-[beta gamma-imido]triphosphate), were ineffective. The metalloendoproteinase inhibitor carbobenzoxy-Gly-Phe-amide reduced the Ca2+ effect by some 50%. Inhibitors of chymotrypsin and the Ca2+ proteinase calpain had no effect. Ca2+ ionophores (A23187 and ionomycin) and the polycations neomycin and polymixin B blocked the Ca2+ response entirely. Neomycin also abolished a Ca2(+)-independent stimulation of ouabain binding by the wasp venom mastoparan. The requirements for increasing the accessibility of ouabain-binding sites are remarkably similar to those for exocytosis in secretory cells, suggesting that oocytes and eggs possess a Ca2(+)-regulated pathway for the plasma membrane insertion of sodium pumps.

show abstract

“…One approach to clarifying such relationships would be to study insulin-mediated glucose transport under conditions where insulin internalization is blocked. We recently characterized a class of agents, chymotrypsin substrate analogues, that efficiently inhibit insulin internalization in isolated rat adipocytes (11). To determine the relationship between insulin internalization and insulin-stimulated glucose transport, we examined the effect of inhibiting insulin internalization with chymotrypsin substrate analogues on insulinstimulated glucose uptake.…”

Section: Chymotrypsin Substrate Analogues Diabetes 36:542-45 1987mentioning

confidence: 99%

“…The cells were filtered through a polystyrene mesh, washed three times, and suspended in Krebs-Ringer phosphate (pH 7.4) containing 1% BSA (KRP/BSA). Adipocyte counts were performed as previously described (11). Measurement of insulin internalization.…”

Section: Materials Porcine Monocomponent Insulin Was Supplied Bymentioning

confidence: 99%

Insulin-Stimulated Glucose Transport and Insulin Internalization Share a Common Postbinding Step in Adipocytes

Jochen

Berhanu

1987

Diabetes

View full text Add to dashboard Cite

We recently demonstrated that chymotrypsin substrate analogues inhibit receptor-mediated insulin internalization in isolated rat adipocytes. In this study, the effect on glucose transport of inhibiting insulin internalization with these agents was examined. Glucose transport was assayed by measuring [3H]-2-deoxyglucose uptake, and internalized insulin was measured after rapidly dissociating surface-bound insulin with an acidic buffer. The chymotrypsin substrate analogue N-acetyl-Tyr ethyl ester inhibited insulin internalization by 85% while increasing surface-bound insulin by 80-110%. Under these conditions, ATP levels were minimally altered, and basal glucose transport was unchanged; however, insulin-stimulated glucose transport was decreased by 86%. The inhibition of insulin-stimulated glucose transport was not overcome by supramaximal concentrations (400 ng/ml) of insulin. When insulin internalization and insulin-stimulated glucose transport were measured in the presence of increasing concentrations of N-acetyl-Tyr ethyl ester (0.1-1 mM), a strong and highly significant correlation (r = .97, P less than .001) was found between inhibition of insulin internalization and inhibition of insulin-stimulated glucose uptake. Fragments of N-acetyl-Tyr ethyl ester that do not inhibit insulin internalization were also without effect on insulin-stimulated glucose transport. In addition to N-acetyl-Tyr ethyl ester, four other chymotrypsin substrate analogues that are effective inhibitors of insulin internalization also markedly inhibited insulin-stimulated glucose transport. These results indicate that insulin internalization and insulin-stimulated glucose transport share a common postbinding step in adipocytes and that this step is inhibitable by chymotrypsin substrate analogues.

show abstract

Chymotrypsin substrate analogues inhibit endocytosis of insulin and insulin receptors in adipocytes.

Cited by 17 publications

References 44 publications

Purification and characterization of a processing protease from rat liver mitochondria.

Purification and characterization of a processing protease from rat liver mitochondria.

Micromolar free calcium exposes ouabain-binding sites in digitonin-permeabilized Xenopus laevis oocytes

Insulin-Stimulated Glucose Transport and Insulin Internalization Share a Common Postbinding Step in Adipocytes

Contact Info

Product

Resources

About