Transplantation of a 17-amino acid alpha-helical DNA-binding domain into an antibody molecule confers sequence-dependent DNA recognition.
Recombinant antibodies capable of sequence-specific interactions with nucleic acids represent a class of DNA- and RNA-binding proteins with potential for broad application in basic research and medicine. We describe the rational design of a DNA-binding antibody, Fab-Ebox, by replacing a variable segment of the immunoglobulin heavy chain with a 17-amino acid domain derived from TFEB, a class B basic helix-loop-helix protein. DNA-binding activity was studied by electrophoretic mobility-shift assays in which Fab-Ebox was shown to form a specific complex with DNA containing the TFEB recognition motif (CACGTG). Similarities were found in the abilities of TFEB and Fab-Ebox to discriminate between oligodeoxyribonucleotides containing altered recognition sequences. Comparable interference of binding by methylation of cytosine residues indicated that Fab-Ebox and TFEB both contact DNA through interactions along the major groove of double-stranded DNA. The results of this study indicate that DNA-binding antibodies of high specificity can be developed by using the modular nature of both immunoglobulins and transcription factors.
McLane
K E
KE
Department of Immunology, Scripps Research Institute, La Jolla, CA 92037, USA.
Burton
D R
DR
Ghazal
P
P
D000595:Amino Acid Sequence; D000906:Antibodies / Q000378:metabolism*; D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D016384:Consensus Sequence; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D018257:Helix-Loop-Helix Motifs; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D016601:RNA-Binding Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000378:metabolism; D013379:Substrate Specificity; D014157:Transcription Factors
D000906:Antibodies; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D017931:DNA Primers; D004268:DNA-Binding Proteins; D007140:Immunoglobulin Fab Fragments; D009363:Neoplasm Proteins; D015345:Oligonucleotide Probes; D016601:RNA-Binding Proteins; D011994:Recombinant Proteins; C064972:TFEB protein, human; D014157:Transcription Factors; D004247:DNA