A Solution NMR Investigation into the Murine Amelogenin Splice-Variant LRAP (Leucine-Rich Amelogenin Protein).
Amelogenins are the dominant proteins present in ameloblasts during the early stages of enamel biomineralization, making up >90% of the matrix protein. Along with the full-length protein there are several splice-variant isoforms of amelogenin present including LRAP (Leucine-Rich Amelogenin Protein), a protein that consists of the first 33 and the last 26 residues of full-length amelogenin. Using solution-state NMR spectroscopy we have assigned the 1H-15N HSQC spectrum of murine LRAP (rp(H)LRAP) in 2% acetic acid at pH 3.0 by making extensive use of previous chemical shift assignments for full-length murine amelogenin (rp(H)M180). This correlation was possible because LRAP, like the full-length protein, is intrinsically disordered under these solution conditions. The major difference between the 1H-15N HSQC spectra of rp(H)M180 and rp(H)LRAP was an additional set of amide resonances for each of the seven non-proline residues between S12* and Y12 at the N-terminus of rp(H)LRAP indicating that the N-terminal region of LRAP exists in two different conformations. Analysis of the proline carbon chemical shifts suggest that the molecular basis for the two states is not a cis-trans isomerization of one or more of the proline residues in the N-terminal region and is likely due to a slow exchange process. As observed with rp(H)M180, residue specific changes in molecular dynamics, manifested by the reduction in intensity and disappearance of 1H-15N HSQC cross peaks, were observed with the addition of NaCl to rp(H)LRAP. These perturbations may signal early events governing supramolecular self-assembly of rp(H)LRAP into nanospheres. However, the different pattern of 1H-15N HSQC cross peak perturbation between rp(H)LRAP and rp(H)M180 in high salt suggest that the termini may behave differently in their respective nanospheres, and perhaps, these differences account for the cell signaling properties attributable to LRAP but not the full-length protein.
- Research Organization:
- Pacific Northwest National Laboratory (PNNL), Richland, WA (US), Environmental Molecular Sciences Laboratory (EMSL)
- Sponsoring Organization:
- USDOE
- DOE Contract Number:
- AC05-76RL01830
- OSTI ID:
- 988613
- Report Number(s):
- PNNL-SA-69686; 19851; 19851a; 400412000
- Journal Information:
- Biochimica et Biophysica Acta--Proteins and Proteomics, 1804(9):1768-1774, Journal Name: Biochimica et Biophysica Acta--Proteins and Proteomics, 1804(9):1768-1774 Journal Issue: 9 Vol. 1804; ISSN 1570-9639
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
99 GENERAL AND MISCELLANEOUS
ACETIC ACID
AMIDES
CARBON
CHEMICAL SHIFT
ENAMELS
Environmental Molecular Sciences Laboratory
ISOMERIZATION
LRAP
intrinsic disorder
nanosphere self-assembly
NMR spectroscopy
PROLINE
PROTEINS
RESIDUES
SPECTRA
SPECTROSCOPY
amelogenesis
biomineralization
cell signalling
enamel