A high-resolution structure of a ligand-bound, soluble form of human monoglyceride lipase (MGL) is presented. The structure highlights a novel conformation of the regulatory lid-domain present in the lipase family as well as the binding mode of a pharmaceutically relevant reversible inhibitor. Analysis of the structure lacking the inhibitor indicates that the closed conformation can accommodate the native substrate 2-arachidonoyl glycerol. A model is proposed in which MGL undergoes conformational and electrostatic changes during the catalytic cycle ultimately resulting in its dissociation from the membrane upon completion of the cycle. In addition, the study outlines a successful approach to transform membrane associated proteins, which tend to aggregate upon purification, into a monomeric and soluble form.

The topological similarity of IL-10 to IFN gamma was totally unexpected, and may be a reflection of the close relationship between the biological effects of these two cytokines. The structure of IL-10 provides insights into the possible modes of conversion of the dimer into monomers, and of putative sites of receptor interactions. The good level of refinement and high resolution of this structure show that the internal disorder often associated with other helical cytokines is not an essential feature of this class of proteins.

The crystal structure of human interleukin-10 (IL-IO) was refined at 1.6 8, resolution against X-ray diffraction data collected at 100 K with the use of synchrotron radiation. Although similar to the IL-IO structure determined previously at room temperature, this low-temperature IL-IO structure contains, in addition, four N-terminal residues, three sulfate anions, and 175 extra water molecules. Whereas the main-chain conformation is preserved, about 30% of the side chains, most of them on the protein surface, assume different conformations. A computer model of a complex of IL-10 with its two soluble receptors was generated based on the topological similarity of IL-IO to interferon-y. The contact region between the cytokine and each receptor shows excellent complementarity of polar and hydrophobic interactions, suggesting that the model is generally correct and should be useful in guiding mutagenesis experiments.

Aberrant activation of matrix metalloproteinases (MMPs) is a common feature of pathological cascades observed in diverse disorders, such as cancer, fibrosis, immune dysregulation, and neurodegenerative diseases. MMP-9, in particular, is highly dynamically regulated in several pathological processes. Development of MMP inhibitors has therefore been an attractive strategy for therapeutic intervention. However, a long history of failed clinical trials has demonstrated that broad-spectrum MMP inhibitors have limited clinical utility, which has spurred the development of inhibitors selective for individual MMPs. Attaining selectivity has been technically challenging because of sequence and structural conservation across the various MMPs. Here, through a biochemical and structural screening paradigm, we have identified JNJ0966, a highly selective compound that inhibited activation of MMP-9 zymogen and subsequent generation of catalytically active enzyme. JNJ0966 had no effect on MMP-1, MMP-2, MMP-3, MMP-9, or MMP-14 catalytic activity and did not inhibit activation of the highly related MMP-2 zymogen. The molecular basis for this activity was characterized as an interaction of JNJ0966 with a structural pocket in proximity to the MMP-9 zymogen cleavage site near Arg-106, which is distinct from the catalytic domain. JNJ0966 was efficacious in reducing disease severity in a mouse experimental autoimmune encephalomyelitis model, demonstrating the viability of this therapeutic approach. This discovery reveals an unprecedented pharmacological approach to MMP inhibition, providing an opportunity to improve selectivity of future clinical drug candidates. Targeting zymogen activation in this manner may also allow for pharmaceutical exploration of other enzymes previously viewed as intractable drug targets. MMPs2 are a family of structurally related zinc-binding proteolytic enzymes that digest extracellular matrix proteins and participate in tissue remodeling and signaling events (1). Currently, ϳ23 MMPs have been identified, comprising secreted and membrane-bound forms, and different family members share some common structural and functional domains and have varying degrees of substrate specificity. Abnormal expression and activation of MMPs has been implicated in the pathogenesis and pathological progression of several different human diseases that are centered in many different tissues in the periphery and central nervous system (2, 3). Initial clinical exploration of synthetic MMP inhibitors was focused on oncology indications, as preventing the breakdown of tissue matrices and barriers was viewed as a potential mechanism to limit tumor metastasis.Despite intensive efforts over many years to develop synthetic MMP inhibitors, only a single MMP inhibitor, Periostat, a tetracycline derivative used in periodontal disease, has progressed into regular clinical use (4). Of the ϳ50 other clinical trials conducted with active site MMP inhibitors, all have failed due to the onset of significant dose-limiting musculoskeletal toxicity ...

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