Practice question

Limited proteolysis of Pol I by subtilisin yields large C-terminal Klenow fragment comprising polymerase and 3' to 5' exonuclease domains, removing N-terminal 5' to 3' exonuclease domain. Klenow retains templated synthesis and proofreading, widely used historically for DNA labeling, second strand cDNA synthesis, and Sanger sequencing before recombinant enzymes available. Loss of 5' to 3' activity prevents unwanted degradation of template during synthesis. Modern Klenow exo-minus version also lacks proofreading for higher efficiency incorporation of modified nucleotides and dideoxy terminators.