When the intermediate filament proteins vimentin and desmin were reacted for a short period of time with the arginine-specific reagent 1,2-cyclohexanedione, the modification had a severe, inhibitory effect on the assembly of intermediate filaments and on the susceptibility of the basic, amino-terminal polypeptide of both proteins to degradation by the intermediate filament-specific, Ca2+-activated proteinase. However, it had only a slightly inhibitory effect on the binding of vimentin and desmin to ribosomal RNA from Ehrlich ascites tumour cells. Since the Ca2+-activated proteinase is very likely to be a trypsin-like enzyme, with a preference for arginyl and lysyl peptide bonds, the results indicate that the arginine residues of the amino-terminal polypeptide of vimentin and desmin are highly essential for filament assembly but largely dispensable for the binding of both proteins to nucleic acids. This was supported by the observation that two breakdown products of vimentin lacking a 5 X 10(3) Mr and an 8 X 10(3) Mr polypeptide from the amino terminus, respectively, did not assemble into intermediate filaments but were still capable of binding to rRNA. Both polypeptides also bound to single-stranded DNA-cellulose under non-denaturing conditions, but passed the affinity column in the presence of 6 M-urea. Thus, the binding of vimentin to nucleic acids appears to be based on two components: a non-specific electrostatic interaction mediated by the positively charged arginine residues of the amino-terminal polypeptide that is insensitive to denaturation by urea, and a specific interaction that is sensitive to denaturation by urea.