The enzymatic basis of processivity in lambda exonuclease.
Lambda exonuclease is a highly processive 5'-->3' exonuclease that degrades double-stranded (ds)DNA. The single-stranded DNA produced by lambda exonuclease is utilized by homologous pairing proteins to carry out homologous recombination. The extensive studies of lambda biology, lambda exonuclease enzymology and the availability of the X-ray crystallographic structure of lambda exonuclease make it a suitable model to dissect the mechanisms of processivity. lambda Exonuclease is a toroidal homotrimeric molecule and this quaternary structure is a recurring theme in proteins engaged in processive reactions in nucleic acid metabolism. We have identified residues in lambda exonuclease involved in recognizing the 5'-phosphate at the ends of broken dsDNA. The preference of lambda exonuclease for a phosphate moiety at 5' dsDNA ends has been established in previous studies; our results indicate that the low activity in the absence of the 5'-phosphate is due to the formation of inert enzyme-substrate complexes. By examining a lambda exonuclease mutant impaired in 5'-phosphate recognition, the significance of catalytic efficiency in modulating the processivity of lambda exonuclease has been elucidated. We propose a model in which processivity of lambda exonuclease is expressed as the net result of competition between pathways that either induce forward translocation or promote reverse translocation and dissociation.
Subramanian
Krithika
K
Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, Miami, FL 33136-6129, USA.
Rutvisuttinunt
Wiriya
W
Scott
Walter
W
Myers
Richard S
RS
D010582:Bacteriophage lambda / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000821:virology; D005090:Exodeoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005816:Genetic Complementation Test; D008958:Models, Molecular; D009154:Mutation; D009841:Oligonucleotides / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D014764:Viral Proteins
D009841:Oligonucleotides; D014764:Viral Proteins; D004247:DNA; D005090:Exodeoxyribonucleases; C043586:exo protein, Bacteriophage lambda