Fig. 1.

Alc1 binds poly(ADP-ribose). (A) Alc1 domain structure. Sequences below diagram show amino acid changes in catalytically inactive Alc1 mutant E175Q and macrodomain mutant D723A. The mutated macrodomain region is compared to the homologous sequence from the AF1521 macrodomain. Snf2N, SNF2 family N-terminal domain; HelicC, Helicase superfamily C-terminal domain, associated with DEXDc-, DEAD-, and DEAH-box proteins; macro, macrodomain. (B) Approximately 100 ng of each protein was incubated with ³²P-labeled PAR in buffer with the indicated NaCl concentrations. PAR binding was detected with a nitrocellulose filter binding assay. (C) Approximately 100 ng (1×) or 200 ng (2×) wild-type or mutant Alc1 was incubated with PAR in buffer containing 0.15 M NaCl. PAR binding was measured as in panel B. F-Alc1, Flag epitope-tagged Alc1; rec, recombinant.