Evolution and Distribution of C7-Cyclitol Synthases in Prokaryotes and Eukaryotes.

2-Epi-5-epi-valiolone synthase (EEVS), a C7-sugar phosphate cyclase (SPC) homologous to 3-dehydroquinate synthase (DHQS), was discovered during studies of the biosynthesis of the C7N-aminocyclitol family of natural products. EEVS was originally thought to be present only in certain actinomycetes, but analyses of genome sequences showed that it is broadly distributed in both prokaryotes and eukaryotes, including vertebrates. Another SPC, desmethyl-4-deoxygadusol synthase (DDGS), was later discovered as being involved in the biosynthesis of mycosporine-like amino acid sunscreen compounds. Current database annotations are quite unreliable, with many EEVSs reported as DHQS, and most DDGSs reported as EEVS, DHQS, or simply hypothetical proteins. Here, we identify sequence features useful for distinguishing these enzymes, report a crystal structure of a representative DDGS showing the high similarity of the EEVS and DDGS enzymes, identify notable active site differences, and demonstrate the importance of two of these active site residues for catalysis by point mutations. Further, we functionally characterized two representatives of a distinct clade equidistant from known EEVS and known DDGS groups and show them to be authentic EEVSs. Moreover, we document and discuss the distribution of genes that encode EEVS and DDGS in various prokaryotes and eukaryotes, including pathogenic bacteria, plant symbionts, nitrogen-fixing bacteria, myxobacteria, cyanobacteria, fungi, stramenopiles, and animals, suggesting their broad potential biological roles in nature.

Clinical and histopathologic features of failed descemet stripping automated endothelial keratoplasty grafts.

PURPOSE: To study the histopathologic features of Descemet stripping automated endothelial keratoplasty (DSAEK) grafts removed after graft failure and to correlate with relevant clinical features. METHODS: Retrospective study of 8 cases in 7 patients who experienced DSAEK graft failure during the surgeons' learning phase. Perioperative clinical findings were correlated with pathologic findings of the explanted graft specimens. Imaging software was used to measure the central and peripheral thickness of each graft. RESULTS: Graft failure occurred in association with graft detachment or excessive surgical manipulation. In 4 cases, the donor tissue detached, including 2 cases with routes for bubble escape. In 3 cases, the graft remained attached but failed to clear. The mean preoperative DSAEK button thickness was 205 +/- 61.8 microm. Average death to preservation time was 8 hours 47 minutes, and average preservation to surgery time was 2 days 22 hours. By light microscopy, explanted donor tissue showed varying degrees of keratocyte degeneration and marked endothelial cell loss. Two specimens with bubble escape had less endothelial loss, and 1 specimen showed epithelial ingrowth at the interface. Repeat keratoplasty (5 DSAEK, 2 penetrating keratoplasty) was successful in 6 of 7 cases. CONCLUSIONS: This series correlates factors that may play a role in DSAEK failure with histopathologic features of explanted DSAEK lenticules. Marked endothelial loss was common in cases with surgical trauma but was less in cases with bubble escape. One specimen showed an epithelial membrane on the stromal interface. Outcomes of repeat DSAEK in these patients seem promising.