Abstract/Details

Characterization of structural changes and large-scale protein dynamics and their influence on metal ion binding by human serum transferrin by ESI MS


2007 2007

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Abstract (summary)

Human serum Transferrin (hTf) is a ∼80 kDa protein, whose function is iron sequestration and transport. The two lobes of the protein (commonly referred to as N- and C-lobes) have a very high degree of structural identity and provide two distinct binding sites for a ferric ion. In addition to iron, serum transferrin also binds a variety of other metals and is believed to provide a route for the in vivo delivery of such metals to cells.

In the present study ESI MS is used to investigate interactions between human serum transferrin and two non-ferrous metals (indium and bismuth), conformational changes upon metal binding, as well as characterize human serum transferrin N-lobe (hTf/2N) global dynamics and functionally important local dynamic events. The In-hTf complex was directly detected by ESI MS; the Bi-hTf complex in solution was established by monitoring the evolution of charge state distributions of transferrin ions upon acid-induced protein unfolding in the presence and in the absence of the metal in solution. The large size of Bi3+ ion is likely to prevent formation of a closed conformation (canonical structure of the holo-protein), resulting in a non-native metal coordination which causes anomalous instability of the transferrin-bismuth complex in the gas phase. The apo-hTf/2N and Fe-hTf/2N were used in hydrogen/deuterium exchange (HDX) measurement for characterizing protein dynamics. In this measurement, back-exchange was corrected for every transferrin N-lobe peptic fragment individually. The results showed that iron binding induce more compact conformation and significant decrease of HDX kinetics around hinge regions and Lys206 which is one amino acid from the dilysine-trigger. However, the changes around iron binding sites are not as significant. Our hypothesis is that instead of having frozen states, transferrin has certain frequency of conformational hopping.

A new method of rapid detection and identification of disulfide-linked peptides in complex proteolytic mixtures was developed utilizing the tendency of collision-activated peptide ions to lose preferentially side chains of select amino acids in the negative ion mode.

Indexing (details)


Subject
Analytical chemistry
Classification
0486: Analytical chemistry
Identifier / keyword
Pure sciences, Bismuth, Indium, Metal-ion binding, Transferrin
Title
Characterization of structural changes and large-scale protein dynamics and their influence on metal ion binding by human serum transferrin by ESI MS
Author
Zhang, Mingxuan
Number of pages
147
Publication year
2007
Degree date
2007
School code
0118
Source
DAI-B 68/02, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
Advisor
Kaltashov, Igor A.
University/institution
University of Massachusetts Amherst
University location
United States -- Massachusetts
Degree
Ph.D.
Source type
Dissertations & Theses
Language
English
Document type
Dissertation/Thesis
Dissertation/thesis number
3254945
ProQuest document ID
304848164
Copyright
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
http://search.proquest.com/docview/304848164
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