The td intron endonuclease I-TevI makes extensive sequence-tolerant contacts across the minor groove of its DNA target.
I-TevI, a double-strand DNA endonuclease encoded by the mobile td intron of phage T4, has specificity for the intronless td allele. Genetic and physical studies indicate that the enzyme makes extensive contacts with its DNA substrate over at least three helical turns and around the circumference of the helix. Remarkably, no single nucleotide within a 48 bp region encompassing this interaction domain is essential for cleavage. Although two subdomains (DI and DII) contain preferred sequences, a third domain (DIII), a primary region of contact with the enzyme, displays much lower sequence preference. While DII and DIII suffice for recognition and binding of I-TevI, all three domains are important for formation of a cleavage-competent complex. Mutational, footprinting and interference studies indicate predominant interactions of I-TevI across the minor groove and phosphate backbone of the DNA. Contacts appear not to be at the single nucleotide level; rather, redundant interactions and/or structural recognition are implied. These unusual properties provide a basis for understanding how I-TevI recognizes T-even phage DNA, which is heavily modified in the major groove. These recognition characteristics may increase the range of natural substrates available to the endonuclease, thereby extending the invasive potential of the mobile intron.
Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany 12201-0509.
M
M
Bryk
S M
SM
Quirk
J E
JE
Mueller
N
N
Loizos
C
C
Lawrence
M
M
Belfort
D001483:Base Sequence; D004279:DNA, Viral / Q000737:chemistry / Q000378:metabolism*; D004706:Endodeoxyribonucleases / Q000235:genetics / Q000378:metabolism*; D004980:Ethane / Q000378:metabolism; D007438:Introns; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D010710:Phosphates / Q000378:metabolism; D013379:Substrate Specificity; D013604:T-Phages / Q000378:metabolism
D004279:DNA, Viral; D010710:Phosphates; D004706:Endodeoxyribonucleases; C065074:I-TEVI endonuclease; D004980:Ethane