Investigation of the interaction between enzyme and inhibitor by the combination of chemical modification, electrospray ionization mass spectrometry and frit-fast atom bombardment liquid chromatography/mass spectrometry.
The interaction between enzyme and its inhibitor, hen egg-white lysozyme and tri-N-acetylglucosamine (NAG3), was studied by the combination of chemical modification, enzymatic digestion, electrospray ionization mass spectrometry and frit-fast atom bombardment liquid chromatography/mass spectrometry. Chemical modification of amino groups, carboxyl groups, and indole groups was carried out independently. In the absence of NAG3, the carboxyl group in Asp 101 was modified by glycinamidation, and the indole group in Trp 62 was modified by Koshland reagent. In the presence of NAG3, the degree of modification of Asp 101 and Trp 62 decreased. It is suggested that Asp 101 and Trp 62 are involved in the interaction with NAG3. The result is consistent with the one obtained by x-ray crystallography. It is indicated that the combination of chemical modification and mass spectrometry may be effective for the investigation of the binding reaction of enzyme to inhibitor and of protein-protein interaction.
Akashi
S
S
Central Research Laboratories, Ajinomoto Co. Inc., Kawasaki, Japan.
Niitsu
U
U
Yuji
R
R
Ide
H
H
Hirayama
K
K
D000588:Amines / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens; D002851:Chromatography, High Pressure Liquid; D002918:Chymotrypsin; D006868:Hydrolysis; D007202:Indicators and Reagents; D013058:Mass Spectrometry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009113:Muramidase / Q000037:antagonists & inhibitors* / Q000737:chemistry; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D016339:Spectrometry, Mass, Fast Atom Bombardment; D014312:Trisaccharides / Q000737:chemistry / Q000494:pharmacology*
D000588:Amines; D007202:Indicators and Reagents; D010446:Peptide Fragments; D014312:Trisaccharides; C019117:N,N',N''-triacetylchitotriose; D009113:Muramidase; D002918:Chymotrypsin