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Structural insights into fibrinogen dynamics using amide hydrogen/deuterium exchange mass spectrometry.


ABSTRACT: We determined the amide hydrogen/deuterium exchange profile of native human fibrinogen under physiologic conditions. After optimization of the quench and proteolysis conditions, more than 1,200 peptides were identified by mass spectrometry, spanning more than 90% of the constituent A?, B?, and ? chain amino acid sequences. The compact central and distal globular regions of fibrinogen were well protected from deuterium exchange, with the exception of the unfolded amino-terminal segments of the A? and B? chains extending from the central region, and the short ? chain "tail" extending from each distal globular region. The triple-helical coiled-coil regions, which bridge the central region to each distal region, were also well protected with the exception of a moderately fast-exchanging area in the middle of each coiled-coil adjacent to the ? chain carbohydrate attachment site. These dynamic regions appear to provide flexibility to the fibrinogen molecule. The ? chain "out loop" contained within each coiled-coil also exchanged rapidly. The ?C domain (A? 392-610) exchanged rapidly, with the exception of a short segment sandwiched between a conserved disulfide linkage in the N-terminal ?C subdomain. This latter finding is consistent with a mostly disordered structure for the ?C domain in native fibrinogen. Analysis of the dysfibrinogen B? 235 Pro/Leu, which exhibits abnormal fibrin structure, revealed enhanced deuterium exchange surrounding the Pro/Leu substitution site as well as in the vicinity of the high affinity calcium binding site and the A knob polymerization pocket within the ?C domain. The implication of these changes with respect to fibrin structure is discussed.

SUBMITTER: Marsh JJ 

PROVIDER: S-EPMC3821761 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Structural insights into fibrinogen dynamics using amide hydrogen/deuterium exchange mass spectrometry.

Marsh James J JJ   Guan Henry S HS   Li Sheng S   Chiles Peter G PG   Tran Danny D   Morris Timothy A TA  

Biochemistry 20130802 32


We determined the amide hydrogen/deuterium exchange profile of native human fibrinogen under physiologic conditions. After optimization of the quench and proteolysis conditions, more than 1,200 peptides were identified by mass spectrometry, spanning more than 90% of the constituent Aα, Bβ, and γ chain amino acid sequences. The compact central and distal globular regions of fibrinogen were well protected from deuterium exchange, with the exception of the unfolded amino-terminal segments of the Aα  ...[more]

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