Reversible associations between human plasma fibronectin and fibrinogen γγ' heterodimer observed by high pressure size exclusion chromatography and dynamic light scattering

Autor: Ayman E.A. Ismail, Ou Wang, Frank M. Fabian, William H. Velander, Yuguo Lei
Rok vydání: 2019
Předmět:
Zdroj: Analytical biochemistry. 598
ISSN: 1096-0309
Popis: We previously reported on a novel fibrin matrix having increased viscoelastic strength derived from human plasma fibronectin (pFN) and γγ′-fibrinogen (γγ′-FI). Here we use high pressure size exclusion chromatography (HPSEC) and dynamic light scattering (DLS) to observe interactions between the linearly extended conformation of γγ′-FI and random coiled pFN. Distinct γγ′-FI:pFN subpopulations were fractionated where each maintained unique retention times when individually reprocessed by HPSEC. The hydrodynamic sizes by HPSEC and DLS for these reprocessed subfractions were intermediate to that of pure γγ′-FI and pFN. SDS-PAGE analysis showed that the majority of these subfractions contained intact γγ′-FI and pFN. Importantly, after disruption and isolation using Gelatin Sepharose affinity chromatography, new complexes rapidly formed between pFN and γγ′-FI when mixed back together. This also occurred in analogous mixing experiments between Des-Aα γγ′-FI and pFN where both Aα-chains are reduced by about 15 kDa due to proteolysis. The reversible complexation observed using HPSEC and DLS was not observed in prior studies using SPR indicating that unrestricted freedom of motion is needed to efficiently form these compact associations. The presence of a γ′ chain, but not the carboxy terminal portions of either Aα chain are needed for complexation phenomena between pFN and γγ′-FI.
Databáze: OpenAIRE
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