Conformation, enzymic activity, and immunochemistry of a lysozyme derivative modified at tryptophan 123 by reaction with 2,3-dioxo-5-indolinesulfonic acid.
Reaction of hen egg-white lysozyme with 2,3-dioxo-5-indolinesulfonic acid (DISA) yielded a homogeneous derivative which was modified at a single tryptophan residue. The modification was located at Trp-123. The absorption spectrum of the derivative showed a new peak in the visible range with lambdamax at 365 nm. In addition, the absorption maximum in the ultraviolet which appears in lysozyme at 280 nm was shifted to 270 nm in the derivative and appreciably enhanced. In ORD measurements, the rotatory behaviors of lysozyme and its derivative were identical at the 233 nm negative minimum and the 199 nm positive extremum. CD measurements gave equal [theta] values for lysozyme and derivative at the two negative ellipticity bands at 208 and 220 nm. Although no conformational differences between lysozyme and derivative were observed by ORD and CD measurements, some changes were detectable by chemical methods. Accessibility to tryptic hydrolysis and susceptibility of the disulfide bonds to reduction were increased in the derivative relative to lysozyme. The lytic activity of the derivative, which retained the same pH optimum as native lysozyme, was greatly (50%) decreased, probably as a result of the slight conformational change. With several antisera to lysozyme, the native protein and its derivative had equal antigenic reactivities. The findings were instrumental in further delineation of an antigenic reactive site in lysozyme.
D000596:Amino Acids / Q000032:analysis; D001665:Binding Sites; D002942:Circular Dichroism; D004529:Egg White; D007211:Indoles; D009113:Muramidase / Q000276:immunology / Q000378:metabolism; D009905:Optical Rotatory Dispersion; D010446:Peptide Fragments / Q000032:analysis; D011233:Precipitin Tests; D011485:Protein Binding; D011487:Protein Conformation; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D014357:Trypsin; D014364:Tryptophan
D000596:Amino Acids; D007211:Indoles; D010446:Peptide Fragments; C011796:2,3-dioxo-5-indolinesulfonic acid; D014364:Tryptophan; D009113:Muramidase; D014357:Trypsin