OBSERVATION OF ALBUMIN RESONANCES IN PROTON NUCLEAR-MAGNETIC-RESONANCE SPECTRA OF HUMAN BLOOD-PLASMA - N-TERMINAL ASSIGNMENTS AIDED BY USE OF MODIFIED RECOMBINANT ALBUMIN

Two-dimensional total shift correlation spectroscopy (TOCSY) and doubl e-quantum-filtered phase-sensitive homonuclear shift-correlated spectr oscopy (DQF-COSY) H-1 NMR spectra are used to assign peaks for about o ne sixth of the amino acids residues of isolated human serum albumin ( 67 kDa) to amino acid types. Sequential assignments are presented for H-1 NMR resonances of the N-terminal residues Asp1, Ala2 and His3 of h uman serum albumin (HSA). These are based on pH-dependent chemical shi fts reflecting the titrating N-terminal NH2 and the His3 imidazole rin g, in addition to DQF-COSY and TOCSY experiments. Studies of variant r ecombinant human albumin with Asp1 deleted, rHA(2-585), aided the assi gnments. The structural nature of the N-and C-termini of HSA are discu ssed and pK(a) values of 7.9 and 6.3 were determined for the N-termina l amino group and His3 imidazole ring, respectively. About 20 spin sys tems for albumin, including those for the N-terminal amino acids, were assigned in H-1 NMR spectra of blood plasma by comparison with isolat ed albumin. Resonances fbr lipids within lipoproteins and also several low molecular mass components can also be assigned in 2D TOCSY H-1 NM R spectra of plasma.