The Escherichia coli OxyR transcription factor senses H2O2 and is activated through the formation of an intramolecular disulfide bond. Here we present the crystal structures of the regulatory domain of OxyR in its reduced and oxidized forms, determined at 2.7 A and 2.3 A resolutions, respectively. In the reduced form, the two redox-active cysteines are separated by approximately 17 A. Disulfide bond formation in the oxidized form results in a significant structural change in the regulatory domain. The structural remodeling, which leads to different oligomeric associations, accounts for the redox-dependent switch in OxyR and provides a novel example of protein regulation by "fold editing" through a reversible disulfide bond formation within a folded domain.

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Plants express numerous calmodulin (CaM) isoforms that exhibit differential activation or inhibition of CaMdependent enzymes in vitro; however, their specificities toward target enzyme/protein binding are uncertain. A random peptide library displaying a 22-mer peptide on a bacteriophage surface was constructed to screen peptides that specifically bind to plant CaM isoforms (soybean calmodulin (ScaM)-1 and SCaM-4 were used in this study) in a Ca 2؉ -dependent manner. The deduced amino acid sequence analyses of the respective 80 phage clones that were independently isolated via affinity panning revealed that SCaM isoforms require distinct amino acid sequences for optimal binding. SCaM-1-binding peptides conform to a 1-5-10 ((FILVW)XXX(FILV) XXXX(FILVW)) motif (where X denotes any amino acid), whereas SCaM-4-binding peptide sequences conform to a 1-8-14 ((FILVW)XXXXXX(FAILVW)XXXXX(FILVW)) motif. These motifs are classified based on the positions of conserved hydrophobic residues. To examine their binding properties further, two representative peptides from each of the SCaM isoform-binding sequences were synthesized and analyzed via gel mobility shift assays, Trp fluorescent spectra analyses, and phosphodiesterase competitive inhibition experiments. The results of these studies suggest that SCaM isoforms possess different binding sequences for optimal target interaction, which therefore may provide a molecular basis for CaM isoform-specific function in plants. Furthermore, the isolated peptide sequences may serve not only as useful CaM-binding sequence references but also as potential reagents for studying CaM isoform-specific function in vivo.

The X-ray crystal structure of inactivated carboxypeptidase A by (2R,3S)-2-benzyl-3,4-epoxybutanoic acid, a pseudomechanism-based inactivator, was determined to show that the carboxylate of Glu-270 at the active site of the enzyme is covalently modified:  the inhibitor is tethered to the carboxylate forming an ester linkage which is brought about by the attack of the carboxylate on the oxirane ring of the inhibitor. Examination of the crystal structure in comparison with that inactivated by its enantiomer, (2S,3R)-2-benzyl-3,4-epoxybutanoic acid, shows that the two inhibitors are bound covalently to the enzyme in a symmetrical fashion. An explanation for the lack of inactivating activity found previously with (2R,3R)- as well as (2S,3S)-2-benzyl-3,4-epoxybutanoic acids was offered.