binary enzyme-substrate (2; throughout the text, the bold numbers refer to species in Fig. The Catalytic Pathway of 1) and enzyme-product (4) complexes have Cytochrome P450cam a t been so characterized (4). Some features of the dioxygen-bound or activated oxygen intermediates, in particular the geometry of the Atomic Resolution six-coordinate low-spin heme, have been deduced from the structure of the ferrous llme ~chlichting,'* Joel Berendzen,' Kelvin Chu,'? Ann M. S t~c k ,~ (FelI+) carbonmonoxy complex (3) of Shelley A. M a v e~,~ David E. enso on,^ Robert M. S~e e t ,~ P450cam (5).However, the binding of carbon Dagmar Ringe,6 Gregory A. ~e t s k o ,~ monoxide to heme is likely to be different in Stephen G. Sligar'~~ a number of important ways from the binding Members o f t h e cytochrome P450 superfamily catalyze t h e addition o f m o -of oxygen ( 6 ) , and regardless, carbon monlecular oxygen t o nonactivated hydrocarbons a t physiological temperature-a oxide is an inhibitor, not a substrate, of reaction t h a t requires high temperature t o proceed i n t h e absence o f a catalyst. P450cam. Hence, the primary evidence for Structures were obtained for three intermediates i n the hydroxylation reaction the structures of the ferrous enzyme-substrate o f camphor b y P450cam w i t h trapping techniques and cryocrystallography. The complex (5), the dioxy intermediate (6), and structure o f t h e ferrous dioxygen adduct o f P450cam was determined w i t h 0.91

The structure of GH5, the globular domain of the linker histone H5, has been solved to 2.5 A resolution by multiwavelength anomalous diffraction on crystals of the selenomethionyl protein. The structure shows a striking similarity to the DNA-binding domain of the catabolite gene activator protein CAP, thereby providing a possible model for the binding of GH5 to DNA.

Bacterial resistance to aminoglycoside antibiotics is almost exclusively accomplished through either phosphorylation, adenylylation, or acetylation of the antibacterial agent. The aminoglycoside kinase, APH(3')-IIIa, catalyzes the phosphorylation of a broad spectrum of aminoglycoside antibiotics. The crystal structure of this enzyme complexed with ADP was determined at 2.2 A. resolution. The three-dimensional fold of APH(3')-IIIa reveals a striking similarity to eukaryotic protein kinases despite a virtually complete lack of sequence homology. Nearly half of the APH(3')-IIIa sequence adopts a conformation identical to that seen in these kinases. Substantial differences are found in the location and conformation of residues presumably responsible for second-substrate specificity. These results indicate that APH(3') enzymes and eukaryotic-type protein kinases share a common ancestor.

Small molecules such as NO, O2, CO or H2 are important biological ligands that bind to metalloproteins to function crucially in processes such as signal transduction, respiration and catalysis. A key issue for understanding the regulation of reaction mechanisms in these systems is whether ligands gain access to the binding sites through specific channels and docking sites, or by random diffusion through the protein matrix. A model system for studying this issue is myoglobin, a simple haem protein. Myoglobin has been studied extensively by spectroscopy, crystallography, computation and theory. It serves as an aid to oxygen diffusion but also binds carbon monoxide, a byproduct of endogenous haem catabolism. Molecular dynamics simulations, random mutagenesis and flash photolysis studies indicate that ligand migration occurs through a limited number of pathways involving docking sites. Here we report the 1.4 A resolution crystal structure of a ligand-binding intermediate in carbonmonoxy myoglobin that may have far-reaching implications for understanding the dynamics of ligand binding and catalysis.

The high mobility group (HMG) chromosomal proteins, which are common to all eukaryotes, bind DNA in a non-sequence-specific fashion to promote chromatin function and gene regulation. They interact directly with nucleosomes and are believed to be modulators of chromatin structure. They are also important in V(D)J recombination and in activating a number of regulators of gene expression, including p53, Hox transcription factors and steroid hormone receptors, by increasing their affinity for DNA. The X-ray crystal structure, at 2.2 A resolution, of the HMG domain of the Drosophila melanogaster protein, HMG-D, bound to DNA provides the first detailed view of a chromosomal HMG domain interacting with linear DNA and reveals the molecular basis of non-sequence-specific DNA recognition. Ser10 forms water-mediated hydrogen bonds to DNA bases, and Val32 with Thr33 partially intercalates the DNA. These two 'sequence-neutral' mechanisms of DNA binding substitute for base-specific hydrogen bonds made by equivalent residues of the sequence-specific HMG domain protein, lymphoid enhancer factor-1. The use of multiple intercalations and water-mediated DNA contacts may prove to be generally important mechanisms by which chromosomal proteins bind to DNA in the minor groove.