The minimal active domain GEF domain. of the mouse Ras exchange factor CDC25Mm was purified to homogeneity from recombinant Escherichia coli culture. The 256 amino acids polypeptide shows high activity in vitro and forms a stable complex with H-ras p21 in absence of guanine nucleotides. Circular dichroism CD. spectra in the far UV region indicate that this domain is highly structured with a high content of a-helix 42%.. Near UV CD spectra evidenced good signal due to phenylalanine and tyrosine while a poor contribution was elicited by the three tryptophan residues contained in this domain. The tryptophan fluorescence signal was scarcely affected by denaturation of the protein or by formation of the binary complex with H-ras p21, suggesting that the Trp residues, which are well conserved in the GEF domain of several Ras-exchange factors, were exposed to the surface of the protein and they are not most probably directly involved in the interaction with Ras proteins. q1998 Elsevier Science B.V
Analysis of the Secondary Structure of the Catalytic Domain of Mouse Ras Exchange Factor CDC25Mm
MONZANI, ENRICO;CASELLA, LUIGI;
1998-01-01
Abstract
The minimal active domain GEF domain. of the mouse Ras exchange factor CDC25Mm was purified to homogeneity from recombinant Escherichia coli culture. The 256 amino acids polypeptide shows high activity in vitro and forms a stable complex with H-ras p21 in absence of guanine nucleotides. Circular dichroism CD. spectra in the far UV region indicate that this domain is highly structured with a high content of a-helix 42%.. Near UV CD spectra evidenced good signal due to phenylalanine and tyrosine while a poor contribution was elicited by the three tryptophan residues contained in this domain. The tryptophan fluorescence signal was scarcely affected by denaturation of the protein or by formation of the binary complex with H-ras p21, suggesting that the Trp residues, which are well conserved in the GEF domain of several Ras-exchange factors, were exposed to the surface of the protein and they are not most probably directly involved in the interaction with Ras proteins. q1998 Elsevier Science B.VI documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
