A simple two-step procedure was developed to identify and characterize genetic variants of apolipoproteins A-I, A-II, and A-IV without the use of ultracentrifugation. The two-step procedure separates the hydrophobic apolipoproteins from the hydrophilic proteins by agarose gel electrophoresis of serum or plasma in the presence of a detergent mixture (Triton X-100 + cetyltrimethylammonium bromide) and then analyzes the isolated, detergent-delipidized apolipoproteins by isoelectric focusing. With this technique, human plasma apolipoprotein A-I focuses as one major and one minor band. Apolipoproteins A-II and A-IV each exhibit single resolved bands. We have demonstrated the potential of this method to detect variant apolipoproteins by finding mutants of apolipoproteins A-I and A-IV, which we have designated apo-A-I-Marburg, apo-A-I-Giessen, and apo-A-IV-Marburg. The method was combined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis for two-dimensional analysis and also with immunochemical procedures to characterize both normal group A apolipoproteins and those from three individuals having variant A-I proteins, apo-A-I-Marburg, apo-A-I-Giessen and apo-A-I-Milano. All three individuals are heterozygotes having normal and variant apolipoprotein A-I. However, the three mutants are not identical but are coded for by different alleles (apo-A-I(Ma), apo-A-I(Gi), and apo-A-I(Mi)). The results show that our method can satisfactorily be used to characterize A apolipoproteins without the use of ultracentrifugation. The identification and characterization of human apolipoprotein mutants will be useful to study structure-function relationships of these proteins.

Genetic variants of group A apolipoproteins. Rapid methods for screening and characterization without ultracentrifugation / G. Utermann, G. Feussner, G. Franceschini, J. Haas, A. Steinmetz. - In: THE JOURNAL OF BIOLOGICAL CHEMISTRY. - ISSN 0021-9258. - 257:1(1982), pp. 501-507.

Genetic variants of group A apolipoproteins. Rapid methods for screening and characterization without ultracentrifugation

G. Franceschini;
1982

Abstract

A simple two-step procedure was developed to identify and characterize genetic variants of apolipoproteins A-I, A-II, and A-IV without the use of ultracentrifugation. The two-step procedure separates the hydrophobic apolipoproteins from the hydrophilic proteins by agarose gel electrophoresis of serum or plasma in the presence of a detergent mixture (Triton X-100 + cetyltrimethylammonium bromide) and then analyzes the isolated, detergent-delipidized apolipoproteins by isoelectric focusing. With this technique, human plasma apolipoprotein A-I focuses as one major and one minor band. Apolipoproteins A-II and A-IV each exhibit single resolved bands. We have demonstrated the potential of this method to detect variant apolipoproteins by finding mutants of apolipoproteins A-I and A-IV, which we have designated apo-A-I-Marburg, apo-A-I-Giessen, and apo-A-IV-Marburg. The method was combined with sodium dodecyl sulfate-polyacrylamide gel electrophoresis for two-dimensional analysis and also with immunochemical procedures to characterize both normal group A apolipoproteins and those from three individuals having variant A-I proteins, apo-A-I-Marburg, apo-A-I-Giessen and apo-A-I-Milano. All three individuals are heterozygotes having normal and variant apolipoprotein A-I. However, the three mutants are not identical but are coded for by different alleles (apo-A-I(Ma), apo-A-I(Gi), and apo-A-I(Mi)). The results show that our method can satisfactorily be used to characterize A apolipoproteins without the use of ultracentrifugation. The identification and characterization of human apolipoprotein mutants will be useful to study structure-function relationships of these proteins.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/182422
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