Probing the sulfhydryl groups of nuclear matrix proteins with 6-iodoacetamidofluorescein.
Rat liver nuclear matrices were reacted with the fluorescent dye 6-iodoacetamidofluorescein and the matrix proteins were then separated by one and two-dimensional polyacrylamide gel electrophoresis. Upon transillumination with U.V. light it was possible to see that several proteins had reacted with the dy, thus indicating the presence of free -SH groups. This labelling technique allowed the detection of a large number of proteins, being several folds more sensitive than conventional Coomassie Blue staining, as demonstrated by two-dimensional electrophoretical separation. If nuclear matrices were treated with reducing agents before being reacted with 6-iodoacetamidofluorescein, the fluorescence increased with about the same intensity in all the protein bands. It is proposed that 6-iodoacetamidofluorescein can be used as a specific and very sensitive probe to study the -SH groups of nuclear matrix proteins.
Martelli
A M
AM
Instituto di Anatomia Umana Normale, Università di Bologna.
Billi
A M
AM
De Marchis
C
C
Manzoli
L
L
Cocco
L
L
D000818:Animals; D015180:Electrophoresis, Gel, Two-Dimensional; D005452:Fluoresceins; D005456:Fluorescent Dyes; D008099:Liver / Q000032:analysis / Q000648:ultrastructure; D008297:Male; D015530:Nuclear Matrix / Q000032:analysis*; D009687:Nuclear Proteins / Q000032:analysis*; D051381:Rats; D011919:Rats, Inbred Strains; D013438:Sulfhydryl Compounds / Q000032:analysis*
D005452:Fluoresceins; D005456:Fluorescent Dyes; D009687:Nuclear Proteins; D013438:Sulfhydryl Compounds; C050046:6-iodoacetamidofluorescein