Disulfide bonds in the SAPA domain of the pulmonary surfactant protein B precursor.

The disulfide bonds formed in the SAPA domain of a recombinant version of the NH-terminal propeptide (SP-B) from the precursor of human pulmonary surfactant protein B (SP-B) were identified through sequential digestion of SP-B with GluC/trypsin or thermolysin/GluC, followed by mass spectrometry (MS) analysis. MS spectra allowed identification of disulfide bonds between Cys-Cys and Cys-Cys, and we propose a disulfide connectivity pattern of 1-3 and 2-4 within the SAPA domain, with the Cys residues numbered according to their position from the N-terminus of the propeptide sequence. The peaks with m/z ∼ 2136 and ∼ 1780 in the MS spectrum of the GluC/trypsin digest were assigned to peptides AWTTSSLACAQGPE and QALQCR linked by Cys-Cys and FWCQSLE and ALGHCLQE linked by Cys-Cys respectively.

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 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.