Fragments of Bovine Serum Albumin

The isolation of a large number of immunochemically reactive fragments with a variety of overlaps and representing various parts of the molecule affords a very effective approach to localizing the antigenic 

---

Complete sequencing of this peptide requires acquisition of additional tandem mass spectra, preferably MS3 fragmentation, of one of the low-mass y-ions. Because the peptide of interest is derived from a biological source, yet another possibility might be the use of sequence databases, similarly to the previous example. Actually, this approach works very well in this case, allowing identification of the peptide of interest as H-LGEYGFQNALIVR-OH, the 421-433 fragment of bovine serum albumin. 

Peters, T., Jr. and Feldhoff, R. C. 1975. Fragments of bovine serum albumin produced by limited proteolysis. Isolation and characterization of peptic fragments. Biochemistry 14, 3384-3391. 

D. Fragmentation of Bovine Serum Albumin by Proteolytic Enzymes... 

That conclusion is supported and extended by the virtually simultaneous publication of Teale and Benjamin (1976). These investigators studied the oxidative regeneration of bovine serum albumin, assaying the extent of refolding immunochemically. Their results showed clearly that some parts of the molecule fold faster than others. Two fragments of albumin were tested for oxidative regeneration in the same way. Substantial return of native structure was seen in both fragments. 

Fig. 11.6. Peptide sequencing by nanoESI-CID-MS/MS from a tryptic digest of bovine serum albumin (BSA) 800 fmol of BSA were used, (a) Eull scan spectrum, (b) fragmentation of the selected doubly charged peptide ion at m/z 740.5. Adapted from Ref. [66] by permission. Nature Publishing Group, 1996.

Haginaka, J. Kanasugi, N. Enantioselectivity of bovine serum albumin-bonded columns produced with isolated protein fragments. J.Chromatogr.A, 1995, 694, 71-80 [chiral also benzoin, clorazepate, fenoprofen, flurbiprofen, ibuprofen, lorazepam, lormetazepam, oxazepam, pranoprofen, temazepam, warfarin]... 

A further support to this view has been afforded from our isolation of another fragment from a tryptic digest of bovine serum albumin (Habeeb and Atassi, 1976a,6). The fragmentation pattern of crystalline native BSA by trypsin was studied (Habeeb and Atassi, 1976a,6) in aqueous solution under various conditions with regard to the yield and size of the... 

Latex particles bearing carbohydrate species were prepared by emulsion copolymerisation of styrene or methyl methacrylate with polymerisable liposacchaiide surfactants. Surface active and mesomorphic properties are discussed. Data are given concerning the adsorption of bovine serum albumin and the covalent binding of antibodies and singlestrand DNA fragments on their surface. 27 refs. 

Amino acid analysis after acid hydrolysis of the pure peptide yields leucine and tyrosine in a 1 1 molar ratio. The peptide material (100 nmoles) is digested with carboxypeptidase A (10 /tg) at pH 8.3 and 37° for 2 hr. The digestion mixture is then passed through a Sephadex G-25 column (1.5 X 30 cm) in 0.1 M ammonia-ammonium acetate at pH 9.2. A complete separation of the peptide fragments is obtained. On the basis of N-terminal and amino acid analysis data on the peptide fragments, and from the known sequence data of bovine serum albumin, it has been concluded that the labeled peptide is the Tyr-Leu-Tyr sequence of residues 137-139 with the fluorescein molecule attached to Tyr-137. 

Urea-induced unfolding of bovine serum albumin and one of its fragments containing domain II+III has been studied by difference spectral and fluorescence emission measurements. The unfolding-refolding curves of both the proteins showed the presence of at least one stable intermediate when the transition was monitored at 288 nm. The presence... 

Figure 6.4. Fragmentation spectrum of a tryptic peptide obtained from bovine serum albumin. Peptide sequence LGEYGFQNALIVR, monoisotopic [M + H]+ = 1479.796, monoisotopic [M+2H]2+ =740.402. Upper panel full scan MS spectrum. Lower panel MS/MS spectrum of a doubly-charged ion at 740.7 m/z with a ladder of y ions, the distances between which correspond to amino acid residues (upper row of letters). A shorter series of b ions is also seen (lower row of letters). See Fig. 6.5 for description of nomenclature. Note the often observed phenomenon where multiply-charged ions lose the charge during fragmentation process and, therefore, have higher m/z values than the original parent ion.

Native bovine serum albumin has the surprising property of catalyzing the decomposition of the Meisenheimer complex 1,1-dihydro-2,4,6-trinitrocylohexadienate. Taylor and Silver (1976) prepared albumin fragments 1-306 and 307-581 and separated them without disulfide reduction. These were called native fragments. Neither of these fragments alone has catalytic activity to decompose the Meisenheimer complex, but when mixed together stoichiometrically, 35%... 

B) O, Anti-bovine serum albumin , anti-C-ffagment and anti-TiM-jM 9, anti-N-fragment. Reprinted, with permission, from Teale and Benjamin (1976). Copyright by the American Society of Biological Chemists, Inc. 

FIGURE 4 Effect of sample preparation on the fragmentation of an rMAb observed in (A) SDS-PAGE and (B) CE-SDS with LIF detection. SDS-PAGE lanes (Lane I) molecular weight standards bovine serum albumin at (Lane 2) 8 ng and (Lane 3) 2 ng (Lane 4) rMAb control after alkylation with (Lane 5) iodoacetic acid and (Lane 6) iodoacetamide. (See color plate 4.)... 

---