Figure 5.

In Vitro Protein–DNA Interaction by EMSA Assays.

(A) Oligonucleotide probe sequences used for EMSA assays. On the basis of the sequences of cbs1 and cbs2, one or two mutated nucleotides were introduced. Bold capital letters represent the consensus sequences, and gray letters indicate mutated nucleotides.

(B) to (D) Retardation of the probes cbs (B), cbs1 (C), and cbs2 (D) by recombinant CYC1C and CYC1D.

(E) CYC1C and CYC1D bind to the probe cbs by forming both homo- and heterodimers.

(F) Mutant probes cbsm1–6 have no or very low binding activity to CYC1C (left) and CYC1D (right) proteins.

(G) The unlabeled oligonucleotides cbs1 and cbs2, rather than cbsm1–6, strongly affect the interaction between the cbs1 probe and the CYC1C (left) or CYC1D (right) protein. −, +, and ++ represent no, onefold, and twofold amounts of recombinant proteins added, respectively.