IQUIFIB   02644
INSTITUTO DE QUIMICA Y FISICOQUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Unidad Ejecutora - UE
artículos
Título:
Experimentally approaching the solvent accessible surface area of a protein. Insights into the acid molten globule of bovine alpha-lactalbumin
Autor/es:
CRAIG PO; GÓMEZ GE; URETA DB; JJ CARAMELO; JM DELFINO
Revista:
JOURNAL OF MOLECULAR BIOLOGY
Editorial:
ACADEMIC PRESS LTD-ELSEVIER SCIENCE LTD
Referencias:
Año: 2009 vol. 394 p. 982 - 993
ISSN:
0022-2836
Resumen:
Each conformational state of a protein is inextricably related to a defined
extent of solvent exposure that plays a key role in protein folding and protein
interactions. However, accurate measurement of the solvent-accessible
surface area (ASA) is difficult for any state other than the native (N) state.
We address this fundamental physicochemical parameter through a new
experimental approach based on the reaction of the photochemical reagent
diazirine (DZN) with the polypeptide chain. By virtue of its size, DZN is a
reasonable molecular mimic of aqueous solvent. Here, we structurally
characterize nonnative states of the paradigmatic protein á-lactalbumin.
Covalent tagging resulting from unspecific methylene (:CH2) reaction allows
one to obtain a global estimate of ASA and to map out solvent accessibility
along the amino acid sequence. By its mild apolar nature, DZN also reveals a
hydrophobic phase in the acid-stabilized state of á-lactalbumin, in which
there is clustering of core residues accessible to the solvent. In a fashion
reminiscent of the N state, this acid-stabilized state also exhibits local regions
where increased :CH2 labeling indicates its nonhomogenous nature, likely
pointing to the existence of packing defects. By contrast, the virtual absence of
a defined long-range organization brings about a featureless labeling pattern
for the unfolded state. Overall, :CH2 labeling emerges as a fruitful technique
that is able to quantify the ASA of the polypeptide chain, thus probing
conformational features such as the outer exposed surface and inner cavities,
as well as revealing the existence of noncompact apolar phases in nonnative
states.
Keywords: diazirine; protein folding; nonnative states; solvent-accessible
surface area; á-lactalbumin