Complete structure elucidation of a globular protein by particle beam liquid chromatography-Fourier transform infrared spectrometry and electrospray liquid chromatography-mass spectrometry. Sequence and conformation of beta-lactoglobulin.

The advantages to the use of both mass spectrometry (MS) and Fourier transform infrared spectrometry (FT-IR) in combination for the structural characterization of the tryptic digest of a model globular protein is demonstrated. HPLC has been interfaced to both spectroscopic techniques and has provided a high degree of structural detail for the target protein. beta-Lactoglobulins A and B were digested with trypsin and chromatographed with narrow-bore, reversed-phase HPLC. As determined by LC-FT-IR spectrometry, the conformation of each form of intact beta-lactoglobulin was randomized upon elution. The particle beam and the electrospray LC-MS interfaces enabled the acquisition of spectra for nanogram injection quantities. The primary structures were determined from the accurate molecular mass determinations of the digest fragments. Infrared spectra confirmed the presence of some amino acid functionalities.