The Diels-Alder reaction is a cornerstone in organic synthesis, forming two carbon-carbon bonds and up to four new stereogenic centers in one step. No naturally occurring enzymes have been shown to catalyze bimolecular Diels-Alder reactions. We describe the de novo computational design and experimental characterization of enzymes catalyzing a bimolecular Diels-Alder reaction with high stereoselectivity and substrate specificity. X-ray crystallography confirms that the structure matches the design for the most active of the enzymes, and binding site substitutions reprogram the substrate specificity. Designed stereoselective catalysts for carbon-carbon bond forming reactions should be broadly useful in synthetic chemistry.

We report a highly regioselective metal-free oxidative cyclization of sulfonamides onto tethered, unactivated alkenes using hypervalent iodine and Brønsted acids. Under these conditions the acid counterion is incorporated into the cyclized products providing an overall aminotrifluoroacetoxylation of the alkene. An unusual preference for endo ring closure is exhibited in contrast to existing exo selective methods. Multiple ring sizes can be formed to access functionalized pyrrolidines, piperidines, and azepanes with a general preference for endo cyclization. A variety of substrate substitution patterns were tolerated to provide nitrogen-containing heterocycles with high regioselectivities and good to excellent diastereoselectivities.

The Morita-Baylis-Hillman reaction forms a carbon-carbon bond between the alpha carbon of a conjugated carbonyl compound and a carbon electrophile. The reaction mechanism involves Michael addition of a nucleophile catalyst at the carbonyl beta carbon, followed by bond formation with the electrophile and catalyst disassociation to release the product. We used Rosetta to design 48 proteins containing active sites predicted to carry out this mechanism, of which two show catalytic activity by mass spectrometry (MS). Substrate labeling measured by MS and site-directed mutagenesis experiments show that the designed active-site residues are responsible for activity, although rate acceleration over background is modest. To characterize the designed proteins, we developed a fluorescence-based screen for intermediate formation in cell lysates, carried out microsecond molecular dynamics simulations, and solved X-ray crystal structures. These data indicate a partially formed active site, and suggest several clear avenues for designing more active catalysts.

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The purpose of this study was to compare the osteogenic potential of a synthetic and a demineralized bone matrix (DBM) putty using a cranial defect model in New Zealand white rabbits. Paired, bilateral critical-size defects (10 mm) were prepared in the frontal bones of 12 rabbits and filled with either OsteoSelect DBM Putty or NovaBone calcium-phosphosilicate putty. At days 43 and 91, 6 rabbits were killed and examined via semiquantitative histology and quantitative histomorphometry. Defects filled with the DBM putty were histologically associated with less inflammation and fibrous tissue in the defect and more new bone than the synthetic counterpart at both time points. Histomorphometric analysis revealed that the defects filled with DBM putty were associated with significantly more bone formation at day 43 (70.7% vs 40.7%, P = 0.043) and at day 91 (70.4% vs 39.9%, P = 0.0044). The amount of residual implant was similar for both test groups at each time point.