This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Amyloid fibrils are composed of abnormally refolded proteins. AFM has demonstrated its capability to elucidate the in-vitro fibril formation process. The fibril formation of amyloid protein such as Amyloid-? and recombinant IgG light?chain has been reported. The model proposed by Ionescu-Zanetti et al shows that the amyloid proteins form ?-sheet structures to become a single filament in the diameter around 2.4 nm. Two or more filaments can intertwine to form larger size protofilbrils or fibrils directly. This AFM project is related to our other amyloid mass spectrometry projects, and focuses on IgG light-chains purified from patient organs and resuspended in solution, as well as the fibrils taken directly from human organs. AL LC fibrils are suspended in buffer and aliquots are taken at different times for AFM analysis under tapping mode. Our preliminary data showed that the rate of fibril formation is very dependent on the incubation conditions, such as pH and stirring. Fibrils were observed at pH 2 with stirring, but not at pH 5.5 and 7.5. The conformation of the amyloid fibrils purified from patient organs were also measured, and the mass spectral characteristics of each of these and their proteolytic digests were also determined. Different experimental conditions are being tested for IgG light-chain fibril formation. The effects of the presence of glycosaminoglycans, e.g., heparin, heparin sulfate, during the fibril formation are also being explored, using highly sensitive and specific methods that have been developed by the Zaia group. A manuscript that reports the results from AFM and EM imaging of fibril growth was recently published in the Journal of Biological Chemistry.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR010888-15
Application #
8365539
Study Section
Special Emphasis Panel (ZRG1-BCMB-H (40))
Project Start
2011-06-01
Project End
2012-08-09
Budget Start
2011-06-01
Budget End
2012-08-31
Support Year
15
Fiscal Year
2011
Total Cost
$15,376
Indirect Cost
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
Hu, Han; Khatri, Kshitij; Zaia, Joseph (2017) Algorithms and design strategies towards automated glycoproteomics analysis. Mass Spectrom Rev 36:475-498
Sethi, Manveen K; Zaia, Joseph (2017) Extracellular matrix proteomics in schizophrenia and Alzheimer's disease. Anal Bioanal Chem 409:379-394
Hu, Han; Khatri, Kshitij; Klein, Joshua et al. (2016) A review of methods for interpretation of glycopeptide tandem mass spectral data. Glycoconj J 33:285-96
Ji, Yuhuan; Bachschmid, Markus M; Costello, Catherine E et al. (2016) S- to N-Palmitoyl Transfer During Proteomic Sample Preparation. J Am Soc Mass Spectrom 27:677-85
Pu, Yi; Ridgeway, Mark E; Glaskin, Rebecca S et al. (2016) Separation and Identification of Isomeric Glycans by Selected Accumulation-Trapped Ion Mobility Spectrometry-Electron Activated Dissociation Tandem Mass Spectrometry. Anal Chem 88:3440-3
Wang, Yun Hwa Walter; Meyer, Rosana D; Bondzie, Philip A et al. (2016) IGPR-1 Is Required for Endothelial Cell-Cell Adhesion and Barrier Function. J Mol Biol 428:5019-5033
Srinivasan, Srimathi; Chitalia, Vipul; Meyer, Rosana D et al. (2015) Hypoxia-induced expression of phosducin-like 3 regulates expression of VEGFR-2 and promotes angiogenesis. Angiogenesis 18:449-62
Yu, Xiang; Sargaeva, Nadezda P; Thompson, Christopher J et al. (2015) In-Source Decay Characterization of Isoaspartate and ?-Peptides. Int J Mass Spectrom 390:101-109
Steinhorn, Benjamin S; Loscalzo, Joseph; Michel, Thomas (2015) Nitroglycerin and Nitric Oxide--A Rondo of Themes in Cardiovascular Therapeutics. N Engl J Med 373:277-80
Walsh, Erica M; Niu, MengMeng; Bergholz, Johann et al. (2015) Nutlin-3 down-regulates retinoblastoma protein expression and inhibits muscle cell differentiation. Biochem Biophys Res Commun 461:293-9

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