Related Experiment Videos
Polymerization defect of fibrinogen Baltimore III due to a gamma Asn308----Ile mutation
1Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, MD.
Abstract:
Fibrinogen Baltimore III, a congenital abnormal fibrinogen with impaired fibrin monomer polymerization, displays a normal gamma-chain and a gamma-variant that has an apparently lower relative molecular weight (mol wt) than normal on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Reverse phase high-performance liquid chromatography (HPLC) analysis of the lysyl endopeptidase digest of the purified gamma-chains of fibrinogen Baltimore III revealed the presence of a peptide that is not found in the digest of the normal fibrinogen gamma-chain. Amino acid sequence analysis of this peptide indicated that the gamma-chain residue 308, asparagine, is replaced by isoleucine. Concanavalin A bound both normal and variant gamma-chains of fibrinogen Baltimore III, indicating that the carbohydrate moiety is not altered and is not responsible for the increase in electrophoretic mobility of the Baltimore III gamma-chain. This study suggests that the integrity of gamma Asn308 is critical for fibrin monomer polymerization, since alteration to either a basic (fibrinogen Kyoto I, Asn----Lys) or hydrophobic (Asn----Ile) residue results in significantly delayed polymerization of fibrinogen to fibrin.
Insights
Fibrinogen Baltimore III, a variant fibrinogen, has a mutation in its gamma-chain (Asn308 to Ile). This alteration impairs fibrin polymerization, highlighting the critical role of gamma Asn308 in clot formation.
Area of Science:
- Biochemistry
- Molecular Biology
- Hematology
Background:
- Congenital dysfibrinogenemia involves abnormal fibrinogen function.
- Fibrinogen Baltimore III exhibits impaired fibrin monomer polymerization.
- The gamma-chain variant shows altered molecular weight on SDS-PAGE.
Purpose of the Study:
- To identify the molecular defect in Fibrinogen Baltimore III.
- To investigate the role of gamma-chain residue 308 in fibrin polymerization.
Main Methods:
- Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE).
- Reverse phase high-performance liquid chromatography (HPLC) of lysyl endopeptidase digests.
- Amino acid sequence analysis.
- Concanavalin A binding assays.
Main Results:
- Identified a unique peptide in the digest of Fibrinogen Baltimore III gamma-chain.
- Determined the mutation: Asn308 replaced by isoleucine (Asn308Ile).
- Confirmed carbohydrate moiety is unaltered and not responsible for electrophoretic mobility changes.
Conclusions:
- The integrity of gamma Asn308 is crucial for normal fibrin monomer polymerization.
- Alterations at residue 308 (e.g., to Lys or Ile) significantly delay fibrinogen to fibrin conversion.