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The hepatic acute-phase proteins alpha 1-antitrypsin and alpha 2-macroglobulin inhibit binding of transferrin to its receptor.

ABSTRACT: Transferrin binding to human placental sites was inhibited by the acute-phase proteins alpha 1-antitrypsin (alpha 1-AT) and alpha 2-macroglobulin (alpha 2-MG), whereas haptoglobin, C-reactive protein and ferritin displayed no such effect. In equilibrium saturation binding assays, the effective acute-phase proteins decreased the apparent affinity of the binding sites for transferrin, but the transferrin binding-site density Bmax. was not significantly changed. For instance, the addition of 30 microM alpha 1-AT increased the KD of transferrin from 8.46 +/- 1.51 nM to 21.6 +/- 3.04 nM; the Bmax. values were 1.17 +/- 0.18 pmol/mg of protein and 1.04 +/- 0.25 pmol/mg of protein respectively. In kinetic studies, alpha 1-AT decreased the association rate constant k+1 of the 125I-transferrin-binding-site complex from 2.18(+/- 0.21) x 10(7) M-1.min-1 to 3.99(+/- 0.18) x 10(6) M-1.min-1. In contrast, the dissociation rate constant k-1 was not changed (0.0948 +/- 0.002 min-1, 0.089 +/- 0.0017 min-1). On isoelectric focusing, no alteration in transferrin protein pattern or shift in isoelectric point was detected in the presence of alpha 1-AT. Inhibition of transferrin binding by the acute-phase proteins alpha 1-AT and alpha 2-MG is competitive. Interestingly, inhibition is already present at physiological concentrations. However, full inhibition is only achieved at concentrations above the normal range, which are attained in acute-phase reactions.

SUBMITTER: Graziadei I

PROVIDER: S-EPMC1132388 | biostudies-other | 1993 Feb

REPOSITORIES: biostudies-other

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Project description:The biosynthesis of rat alpha 1-inhibitor3, a negative acute-phase reactant specifically found in rodents, was studied in vitro in a cell-free translation system from rabbit reticulocytes, in rat hepatocyte primary cultures and in vivo by immunocytochemistry using normal and turpentine-injected rats. By sucrose-gradient centrifugation and subsequent translation of the fractionated RNA in vitro it was found that the mRNA coding for alpha 1-inhibitor3 exhibited a size of about 28S. For the alpha 1-inhibitor3 translated in vitro an apparent Mr of 155,000 was determined. A continuous decrease in the level of alpha 1-inhibitor3 in serum during experimental inflammation induced by turpentine injection was demonstrated by means of quantitative 'rocket' immunoelectrophoresis. This result agrees with the observation by immunocytochemistry of a drastic decrease in alpha 1-inhibitor3 levels in hepatocytes 24 h after turpentine injection. At that time alpha 1-inhibitor3 is mainly located in the Golgi apparatus, whereas it is also present in the membranes of the rough and smooth endoplasmic reticulum when normal liver is used. All hepatocytes, but no other hepatic cells, contain alpha 1-inhibitor3. When hepatocyte primary cultures were labelled with [35S]methionine and alpha 1-inhibitor3 was immunoprecipitated from the hepatocyte medium and the supernatant of homogenized cells, two different forms of alpha 1-inhibitor3 were found. The intracellular form of alpha 1-inhibitor3, with an apparent Mr of 173,000, is characterized by oligosaccharide side chains of the high-mannose type. The form of alpha 1-inhibitor3 in the medium exhibited an Mr of 186,000 and carried carbohydrate side chains of the complex type. After labelling hepatocytes with radioactive sugars, [3H]mannose was found in both forms of alpha 1-inhibitor3, whereas [3H]fucose and [3H]galactose were incorporated only into the form found in the medium. In the presence of tunicamycin an unglycosylated alpha 1-inhibitor3 with an apparent Mr of 154,000 was found in cells and in the medium. In a pulse-chase experiment it was shown that inhibition of glycosylation by tunicamycin resulted in a marked delay of secretion of alpha 1-inhibitor3. Thus the oligosaccharide side chains of alpha 1-inhibitor3 play an important role during its transport into the medium.

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The hepatic acute-phase proteins alpha 1-antitrypsin and alpha 2-macroglobulin inhibit binding of transferrin to its receptor.

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