Conformational stability of the NH2-Terminal propeptide of the precursor of pulmonary surfactant protein SP-B

Assembly of pulmonary surfactant lipid-protein complexes depends on conformational changes coupled with proteolytic maturation of proSP-B, the precursor of pulmonary surfactant protein B (SP-B), along the surfactant biogenesis pathway in pneumocytes. Conformational destabilization of the N-terminal...

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Bibliographic Details
Authors: Bañares Hidalgo, Ángeles, Pérez-Gil, Jesús, Estrada, Pilar
Format: article
Publication Date:2016
Country:España
Institution:Universidad Complutense de Madrid (UCM)
Repository:Docta Complutense
Language:English
OAI Identifier:oai:docta.ucm.es:20.500.14352/23387
Online Access:https://hdl.handle.net/20.500.14352/23387
Access Level:Open access
Keyword:577.112
Urea
Oligomers
Fluorescence
Sedimentation
Surfactants
Protein structure
Thermodynamics
Disulfide bonds
Bioquímica (Biología)
2302 Bioquímica
Description
Summary:Assembly of pulmonary surfactant lipid-protein complexes depends on conformational changes coupled with proteolytic maturation of proSP-B, the precursor of pulmonary surfactant protein B (SP-B), along the surfactant biogenesis pathway in pneumocytes. Conformational destabilization of the N-terminal propeptide of proSP-B (SP-BN) triggers exposure of the mature SP-B domain for insertion into surfactant lipids. We have studied the conformational stability during GdmCl- or urea-promoted unfolding of SP-BN with trp fluorescence and circular dichroism spectroscopies. Binding of the intermediate states to bis-ANS suggests their molten globule-like character. ΔG0 H2O was ~ 12.7 kJ mol-1 either with urea or GdmCl. None of the thermal transitions of SP-BN detected by CD correspond to protein unfolding. Differential scanning calorimetry of SP-BN evidenced two endothermic peaks involved in oligomer dissociation as confirmed with 2 M urea. Ionic strength was relevant since at 150 mM NaCl, the process originating the endotherm at the highest temperature was irreversible (Tm2 = 108.5°C) with an activation energy of 703.8 kJ mol-1. At 500 mM NaCl the process became reversible (Tm2 = 114.4°C) and data were fitted to the Non-two States model with two subpeaks. No free thiols in the propeptide could be titrated by DTNB with or without 5.7 M GdmCl, indicating disulfide bonds establishment.

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