Abstract
Mammalian GATA transcription factors are expressed in various tissues in a temporally regulated manner. The prototypic member, GATA-1, is required for normal erythroid, megakaryocytic, and mast cell development. This family of DNA-binding proteins recognizes a consensus (A/T)GATA(A/G) motif and possesses homologous DNA binding domains consisting of two zinc fingers. The C-terminal finger of GATA-1 recognizes the consensus motif with nanomolar affinities, whereas the N-terminal finger shows a binding preference for a GATC motif, albeit with much reduced affinity (Kd approximately microm). The N-terminal finger of GATA-2 also shows a preference for an AGATCT binding site, with an increased affinity attributed to N- and C-terminal flanking basic residues (Kd approximately nm). To understand the differences in the binding specificities of the N- and C-terminal zinc fingers of GATA-1, we have constructed a series of swapped domain peptides. We show that the specificity for AGATAA over AGATCT arises from the C-terminal non-finger basic domain. Thus, the N-terminal finger binds preferentially to AGATAA once appended to the C-terminal arm of the C-terminal finger. We further show that this specificity arises from the highly conserved QTRNRK residues. The converse is, however, untrue in the case of the C-terminal finger; swapping of QTRNRK with the corresponding LVSKRA does not switch the DNA binding specificity from AGATAA to AGATCT. These results highlight the important role of residues adjacent to the CXXCX17CNAC zinc finger motif (i.e. non-finger residues) in the specific recognition of DNA residues.
Highlights
Unlike the GATA-2 N-finger, this peptide shows a distinct high affinity preference for GATA motifs (Table III), in a manner similar to the analogous peptide NC1. These results demonstrate that fusion of the C-terminal basic arm of GATA-1 to the N-terminal zinc fingers of GATA-1 or -2 alters their binding specificity from GATC to that observed for the full-length protein or C-terminal zinc finger
The N-terminal zinc finger binds weakly to DNA with a preference for GATC sites, whereas the C-terminal zinc finger binds to GATA sites with high affinity [4, 9]
The N- and C-terminal core finger residues of GATA-1 recognize the GAT motif on DNA, as expected from their homology, a result confirmed by selection experiments with the GATA-2 N-terminal finger, which shares the same core finger residues as the N-terminal finger of GATA-1
Summary
The converse is, untrue in the case of the C-terminal finger; swapping of QTRNRK with the corresponding LVSKRA does not switch the DNA binding specificity from AGATAA to AGATCT These results highlight the important role of residues adjacent to the CXXCX17CNAC zinc finger motif (i.e. non-finger residues) in the specific recognition of DNA residues. Solution NMR studies of the complex formed between the C-terminal finger of cGATA-1 and the AGATAA DNA sequence highlight the role that the C-terminal basic arm plays in the high affinity recognition process [11]. Replacement of the QTRNRK residues with LVSKRA in the C-terminal zinc finger does not result in the loss of specificity for the GATAA motif; rather equal specificities are noted for both GATC and GATA motifs
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
