THE LAMBDA-TERMINASE ENZYME MEASURES THE POINT OF ITS ENDONUCLEOLYTICATTACK 47+ -2BP AWAY FROM ITS SITE OF SPECIFIC DNA-BINDING, THE R-SITE/

lambda terminase is an ATP-interactive, site-specific endonuclease com prising the products of lambda genes Nu1 and A. Terminase binds to cos , at the junction of two chromosomes in a concatemer, catalyzes cos cl eavage and initiates the packaging of lambda DNA into proheads. cos co nsists of a nicking domain, cosN, where terminase cleaves to regenerat e the 12 nucleotide cohesive ends of mature lambda chromosomes and a b inding domain, cosB, where terminase binds to 16 bp repeat sequences c alled R3, R2 and R1. Evidence is presented that terminase is a single- strand endonuclease that can nick DNA by one of two mechanisms, both o f which require ATP. (i) When bound to any R site, terminase nicks the strand which, within that R site, is purine-rich; the position of thi s nick is 47 +/- 2 nucleotides away from the mid-point of that R site, measured in the 3' direction; (ii) enzymes that are not bound to R si tes nick DNA within certain specific sequences that resemble cosN half sites. These two modes of action are nicely combined for the R3-bound protomer that nicks the bottom strand at position N1 in cosN since th e interval between N1 and the R3 midpoint is 47 nucleotides. Within co sN, the bottom and top strand nicks are generated by a rigid protein c ouple with a 2-fold rotational symmetry. The location of both of these nicks, however, is gauged asymmetrically from R3, 47 nucleotides away , Again, R1 and R2 are separated by 47 bp and orient bound protomers t owards each other but, unless the DNA between these R sites is lengthe ned, the enzymes do not nick, indicating an inhibitory gpA-gpNu1 appos ition.