A single amino acid substitution in a histidine-transport protein drastically alters its mobility in sodium dodecyl sulfate-polyacrylamide gel electrophoresis

Biochemistry ◽  
1979 ◽  
Vol 18 (19) ◽  
pp. 4159-4165 ◽  
Author(s):  
Dale Noel ◽  
Kishiko Nikaido ◽  
Giovanna Ferro-Luzzi Ames
Keyword(s):  
Amino Acid ◽  
Sodium Dodecyl Sulfate ◽  
Amino Acid Substitution ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Transport Protein ◽  
Polyacrylamide Gel ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid

scholarly journals Antithrombin III Budapest: a single amino acid substitution (429Pro to Leu) in a region highly conserved in the serpin family

Blood ◽  
1992 ◽  
Vol 79 (5) ◽  
pp. 1206-1212 ◽  
Author(s):  
RJ Olds ◽  
DA Lane ◽  
R Caso ◽  
M Panico ◽  
HR Morris ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Antithrombin Iii ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Sds Page ◽  
Additional Support ◽  
Variant Protein

Abstract Antithrombin III (AT) is a major plasma serine protease inhibitor and a member of the serpin family of proteins. We have characterized the molecular and genetic basis of AT Budapest, an inherited variant of AT that is associated with thrombotic disease in affected family members. A single amino acid substitution, 429Pro to Leu, was identified, occurring in a region of the molecule that is highly conserved in members of the serpin family. Two forms of variant protein were present in approximately equal amounts in the plasma of the propositus, who is homozygous for the mutation. One form, which had apparently normal Mr, bound heparin strongly and retained some residual thrombin inhibitory activity. The other form had only weak heparin affinity and no antiproteinase activity, and had slightly decreased mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under nonreducing conditions; this normalized in the presence of a reducing agent, suggesting it was caused by a change in conformation. Additional support for a difference in conformation of the two forms of variant was provided by the finding that the fraction that bound heparin- Sepharose was recognized by a monoclonal antibody raised against normal AT, whereas the weak-affinity fraction was not.

scholarly journals Antithrombin III Budapest: a single amino acid substitution (429Pro to Leu) in a region highly conserved in the serpin family

Blood ◽  
1992 ◽  
Vol 79 (5) ◽  
pp. 1206-1212 ◽  
Author(s):  
RJ Olds ◽  
DA Lane ◽  
R Caso ◽  
M Panico ◽  
HR Morris ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Substitution ◽  
Antithrombin Iii ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Single Amino Acid Substitution ◽  
Single Amino Acid ◽  
Sds Page ◽  
Additional Support ◽  
Variant Protein

Antithrombin III (AT) is a major plasma serine protease inhibitor and a member of the serpin family of proteins. We have characterized the molecular and genetic basis of AT Budapest, an inherited variant of AT that is associated with thrombotic disease in affected family members. A single amino acid substitution, 429Pro to Leu, was identified, occurring in a region of the molecule that is highly conserved in members of the serpin family. Two forms of variant protein were present in approximately equal amounts in the plasma of the propositus, who is homozygous for the mutation. One form, which had apparently normal Mr, bound heparin strongly and retained some residual thrombin inhibitory activity. The other form had only weak heparin affinity and no antiproteinase activity, and had slightly decreased mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under nonreducing conditions; this normalized in the presence of a reducing agent, suggesting it was caused by a change in conformation. Additional support for a difference in conformation of the two forms of variant was provided by the finding that the fraction that bound heparin- Sepharose was recognized by a monoclonal antibody raised against normal AT, whereas the weak-affinity fraction was not.

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