A comprehensive sequence alignment of 64 members of the family of matrix metalloproteinases (MMPs) for the entire sequences, and subsequently the catalytic and the hemopexin-like domains, have been performed. The 64 MMPs were selected from plants, invertebrates, and vertebrates. The analyses disclosed that as many as 23 distinct subfamilies of these proteins are known to exist. Information from the sequence alignments was correlated with structures, both crystallographic as well as computational, of the catalytic domains for the 23 representative members of the MMP family. A survey of the metal binding sites and two loops containing variable sequences of amino acids, which are important for substrate interactions, are discussed. The collective data support the proposal that the assembly of the domains into multidomain enzymes was likely to be an early evolutionary event. This was followed by diversification, perhaps in parallel among the MMPs, in a subsequent evolutionary time scale. Analysis indicates that a retrograde structure simplification may have accounted for the evolution of MMPs with simple domain constituents, such as matrilysin, from the larger and more elaborate enzymes.

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The structural basis of the outer membrane permeability for the bacterium Escherichia coli is studied by atomic force microscopy (AFM) in conjunction with biochemical treatment and analysis. The surface of the bacterium is visualized with unprecedented detail at 50 and 5 Å lateral and vertical resolutions, respectively. The AFM images reveal that the outer membrane of native E. coli exhibits protrusions that correspond to patches of lipopolysaccharide (LPS) containing hundreds to thousands of LPS molecules. The packing of the nearest neighbor patches is tight, and as such the LPS layer provides an effective permeability barrier for the Gram-negative bacteria. Treatment with 50 mM EDTA results in the release of LPS molecules from the boundaries of some patches. Further metal depletion produces many irregularly shaped pits at the outer membrane, which is the consequence of progressive release of LPS molecules and membrane proteins. The EDTA-treated cells were analyzed for metal content and for their reactivities toward lysozyme and antibodies specific for LPS. The experiments collectively indicate that the metal depletion procedure did not remove all the LPS molecules despite a dramatic decrease in the metal content. The remaining LPS molecules are present outside the pits, whereas the bottom of the pits is devoid of these molecules. This new structure for the outer membrane exhibits higher permeability than that for the native cells.

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Bacteria resistant to antibiotics are being selected in a relatively short time, and cases of infections resistant to treatment by all known antibiotics are being identified at alarming rates. The primary mechanism for resistance to -lactam antibiotics is the catalytic function of -lactamases. However, imipenem (a -lactam) resists the action of most -lactamases and is virtually the last effective agent against the vancomycin-resistant Gram-positive bacteria, as well as against multiple antibiotic-resistant Gram-negative organisms. Here, we report the crystal structure, to 1.8 Å resolution, of an acyl-enzyme intermediate for imipenem bound to the TEM-1 -lactamase from Escherichia coli, the parent enzyme of 67 clinical variants. The structure indicates an unprecedented conformational change for the complex which accounts for the ability of this antibiotic to resist hydrolytic deactivation by -lactamases. Computational molecular dynamics underscored the importance of the motion of the acyl-enzyme intermediate, which may be a general feature for catalysis by these enzymes.