RNA recognition by transcriptional antiterminators of the BglG/SacY family: Functional and structural comparison of the CAT domain from SacY and LicT

Transcriptional antiterminators of the BglG/SacY family are regulatory prot eins that mediate the induction of sugar metabolizing operons in Gram-posit ive and Gram-negative bacteria. Upon activation, these proteins bind to spe cific targets in nascent mRNAs, thereby preventing abortive dissociation of the RNA polymerase from the DNA template. We have previously characterized the RNA-binding domain of SacY from Bacillus subtilis and determined its t hree-dimensional structure by both NMR and crystallography. hn the present study, we have characterized the paralogous domain from LicT and we present the first structural comparison between two BglG/SacY family members. Simi lar to SacY, the RNA-binding activity of LicT is contained within the 56 N- terminal amino acid residue fragment corresponding to the so-called co-anti terminator (CAT) domain. Surface plasmon resonance affinity measurements sh ow that, compared to SacY-CAT, LicT-CAT binds more tightly and more specifi cally to its cognate RNA target, with a K-D value of about 10(-8) M. The cr ystal structure of LicT-CAT has been determined at 1.8 Angstrom resolution and compared to that of SacY-CAT. Both molecules fold as symmetrical dimers , each monomer comprising a four-stranded antiparallel beta-sheet that stac ks against the beta-sheet of the other monomer in a very conserved manner. Comparison of the proposed RNA-binding surfaces shows that many of the cons erved atoms concentrate in a central region across one face of the CAT dime r, whereas variable elements are mostly found at the edges. Interestingly, the electrostatic potential maps calculated for the two molecules are quite different, except for the core of the RNA-binding site, which appears esse ntially neutral in both structures. (C) 1999 Academic Press.