FIG. 6.

Cross-linking of PII to NRII mutants. PII (E44C/C73S)-TFPAM-3 (9 μM) was incubated with the indicated NRII protein (4.5 μM) in reaction mixtures containing 50 μM 2-ketoglutarate and 0.5 mM ATP as described in Materials and Methods. Cross-linking was initiated by exposure to UV light for 20 min as indicated. (A) Reactions were stopped by addition of SDS gel-loading buffer, and the mixtures were analyzed on SDS-15% polyacrylamide gels. Molecular mass markers are indicated in kilodaltons. (B) Alternatively, reactions were stopped by addition of 120 mM glycine (pH 9.0) and the mixtures were analyzed on 10% nondenaturing polyacrylamide gels. For both panels, the gels were stained with Coomassie brilliant blue R-250.