The broad long term objectives are twofold. One is to develop methods that will extend the power of solution NMR beyond the study of native globular proteins to the study of nonnative and nonglobular proteins. The second objective is to use NMR to understand protein folding mechanisms by studying structures of partially folded proteins, and by assessing the effect of amino acid substitutions on structure, stability and folding kinetics. These studies will form the basis for using NMR to understand how interruptions in the Gly-X-Y pattern, found in collagen diseases like Osteogenesis Imperfecta and Ehlers Danlos Syndrome, can result in serious disease.
The first aim i s to characterize the partially folded state of guinea pig alpha-lactalbumin to elucidate the nature of protein folding intermediates. More specifically, we wish to learn which regions of the molten globule state contain regions of secondary structure and whether tertiary interactions are important in stabilizing these regions of secondary structure. We will characterize the partially folded state by 1H NMR methods, and by isotope labelling and heteronuclear 2D and 3D NMR experiments. Key mutants will be made to assess the effects of sequence change on secondary structure and tertiary interactions of the partially folded state.
The second aim i s to obtain, for the first time, individual residue assignments and the NMR solution structure of triple helical peptides, to determine the role of individual amino acids in stabilizing the triple helix and to understand how key residues direct protein folding. We will examine the effects, by 1D NMR, of (Gly-X-Y) sequence changes on the amount of triple helix formed, and on the kinetics and thermodynamics of folding. To obtain the solution structure we will first design synthetic triple helical peptides to facilitate the spin system identification process. Then we propose 1H NMR experiments as well as heteronuclear NMR experiments that should allow us to perform sequential resonance assignments, and distinguish inter from intra strand NOE's.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM045302-07
Application #
2459420
Study Section
Special Emphasis Panel (ZRG3-BBCA (01))
Project Start
1991-08-01
Project End
2000-07-31
Budget Start
1997-08-01
Budget End
1998-07-31
Support Year
7
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Rutgers University
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
038633251
City
New Brunswick
State
NJ
Country
United States
Zip Code
08901
Zhu, Jie; Hoop, Cody L; Case, David A et al. (2018) Cryptic binding sites become accessible through surface reconstruction of the type I collagen fibril. Sci Rep 8:16646
Hoop, Cody L; Zhu, Jie; Nunes, Ana Monica et al. (2017) Revealing Accessibility of Cryptic Protein Binding Sites within the Functional Collagen Fibril. Biomolecules 7:
Nunes, Ana Monica; Zhu, Jie; Jezioro, Jacqueline et al. (2016) Intrinsic local destabilization of the C-terminus predisposes integrin ?1 I domain to a conformational switch induced by collagen binding. Protein Sci 25:1672-81
Fu, Iwen; Case, David A; Baum, Jean (2015) Dynamic Water-Mediated Hydrogen Bonding in a Collagen Model Peptide. Biochemistry 54:6029-37
Xiao, Jianxi; Sun, Xiuxia; Madhan, Balaraman et al. (2015) NMR studies demonstrate a unique AAB composition and chain register for a heterotrimeric type IV collagen model peptide containing a natural interruption site. J Biol Chem 290:24201-9
Xiao, Jianxi; Yang, Zhangfu; Sun, Xiuxia et al. (2015) Local amino acid sequence patterns dominate the heterogeneous phenotype for the collagen connective tissue disease Osteogenesis Imperfecta resulting from Gly mutations. J Struct Biol 192:127-37
Kim, Seho; Wu, Kuen-Phon; Baum, Jean (2013) Fast hydrogen exchange affects ýýýýýN relaxation measurements in intrinsically disordered proteins. J Biomol NMR 55:249-56
Parmar, Avanish S; Nunes, Ana Monica; Baum, Jean et al. (2012) A peptide study of the relationship between the collagen triple-helix and amyloid. Biopolymers 97:795-806
Kang, Lijuan; Wu, Kuen-Phon; Vendruscolo, Michele et al. (2011) The A53T mutation is key in defining the differences in the aggregation kinetics of human and mouse ?-synuclein. J Am Chem Soc 133:13465-70
Xiao, Jianxi; Cheng, Haiming; Silva, Teresita et al. (2011) Osteogenesis imperfecta missense mutations in collagen: structural consequences of a glycine to alanine replacement at a highly charged site. Biochemistry 50:10771-80

Showing the most recent 10 out of 21 publications