Heterogeneity of protein labeling with a fluorogenic reagent, 3-(2-furoyl)quinoline-2-carboxaldehyde.
Fluorogenic reagents are used for protein labeling when high-sensitivity fluorescence detection is required. Similar to traditional labeling with activated fluorescent dyes, such as fluorescein isothiocyanate, a fluorogenic reaction is expected to change the physical-chemical properties of proteins. Knowledge of these changes may be essential for efficient separation and identification of labeled proteins. Here we studied the effect of labeling of myoglobin with a fluorogenic reagent on the acid-base properties of the protein. The fluorogenic reagent used was 3-(2-furoyl)quinoline-2-carboxaldehyde (FQ). In slab-gel isoelectric focusing, we found that the labeling reaction generated at least six species with pI values lower than that of non-labeled myoglobin. These species can be identified as products of progressive labeling of myoglobin with one to six FQ molecules. The same series of FQ-labeled species were observed when the reaction products were analyzed by capillary zone electrophoresis. The comparison of experimental and theoretical pI values allowed us to elucidate the labeling pattern--the number of FQ molecules corresponding to each labeled product detected by isoelectric focusing.
Stoyanov
Alexander V
AV
Department of Chemistry, York University, 4700 Keele Street, Toronto, Ontario, Canada M3J 1P3.
Ahmadzadeh
Hossein
H
Krylov
Sergey N
SN
D019075:Electrophoresis, Capillary / Q000379:methods; D005456:Fluorescent Dyes / Q000737:chemistry*; D005663:Furans / Q000737:chemistry*; D007525:Isoelectric Focusing; D011506:Proteins / Q000737:chemistry*; D011804:Quinolines / Q000737:chemistry*
D005456:Fluorescent Dyes; D005663:Furans; D011506:Proteins; D011804:Quinolines; C063756:3-(2-furoyl)quinoline-2-carbaldehyde