Protein–Nucleic Acid Interaction: Major Groove Recognition Determinants

Nucleic acids and proteins are two of the most important biomolecules in any living organism, with the former carrying genetic information and the latter executing and regulating the life processes. Protein–nucleic acid interactions therefore play a crucial role in central biological processes, ranging from the mechanism of replication, transcription and recombination to enzymatic events utilizing nucleic acids as substrates. For these reasons the biochemical and structural bases of the protein–DNA recognition process has been a field of intense study. X-ray crystallography, along with NMR and many other chemical and physical methods are applied to analyse how protein and DNA interact with each other. In late 1970s the footprinting technique was evolved to detect protein–DNA binding specificity. The first X-ray crystallographic study of a protein–DNA complex was reported in 1984, and about ten structures were available by the end of the 1980s. The last decade of the twentieth century saw an explosive growth in high-resolution structures of protein–DNA complexes. Over 240 structures of protein–DNA complexes have been determined, of which about 220 were obtained by X-ray crystallography and about 20 by high-resolution NMR studies. Protein–DNA interactions in these complexes have been extensively documented for individual structures and the literature on the subject has been reviewed (Steitz, 1990; Luisi, 1995; Sundaralingam and Burkhart, 1997; Luscombe et al., 2000). The structures of protein–DNA complexes are available in the Nucleic Acid Database (NDB; [http:// ndbserver.rutgers.edu/NDB/index.html]) and the Protein Data Bank (PDB; [http://www.rcsb.org/pdb]). According to their functions, DNA–protein complexes can be grouped into several classes, including polymerases, transcription factors, nucleases and other enzymes and structural proteins. The first group comprises DNA/RNA polymerase interactions, where the protein uses DNA as the template and individual nucleotides as the substrates to synthesize the DNA or RNA polymers. The second large group contains the proteins for gene regulation, most of which are transcription factors. These proteins recognize specific DNA sites and are usually highly sensitive to the DNA sequence. The third class is the nuclease interaction, including endonuclease and exonuclease. The enzymes find the cutting site on the DNA and hydrolyse the P–O bonds. The DNA recombinase also has a similar function. In addition, many proteins are involved in other processes like DNA base modification and the chromosomal proteins provide skeletons for chromatin. Even before any structural work was available, exploration had already begun of the central question of protein– DNA interaction – how the DNA sequence is recognized by a protein. Hypotheses were made about the interaction between the amino acid side-chain of the protein and the nucleic acid bases (Sundaralingam, 1975). It was predicted that an array of possible specific hydrogenbonding interactions between nucleic acid bases and proteins could exist, including both main-chain and sidechain atoms, particularly the side-chains of arginine, asparagine and glutamine for specific contacts with most bases in the DNA, and especially guanines in the major and the minor grooves (Sundaralingam, 1975). These aspects were also discussed by Seeman et al. (1976), who in addition showed that it could be possible to discriminate all potential base pairs (C.G,G.C, A.U and U.A) and identify a specific sequence by examining their hydrogen-bonding sites in the major groove. From studies on DNA-binding protein structures. Matthews and colleagues theorized that the cro repressor protein (a helix) was bound to the major groove of DNA (Ohlendorf et al., 1982). This hypothesis was proved to be correct and the a helix binding in the Article