Michael Gmachl

SNAREpins: minimal machinery for membrane fusion.

Recombinant v- and t-SNARE proteins reconstituted into separate lipid bilayer vesicles assemble into SNAREpins-SNARE complexes linking two membranes. This leads to spontaneous fusion of the docked membranes at physiological temperature. Docked unfused intermediates can accumulate at lower temperatures and can fuse when brought to physiological temperature. A supply of unassembled v- and t-SNAREs is needed for these intermediates to form, but not for the fusion that follows. These data imply that SNAREpins are the minimal machinery for cellular membrane fusion.

The human sperm protein PH-20 has hyaluronidase activity

The PH‐20 protein present on the membrane of guinea pig sperm was characterized using a monoclonal antibody [(1991) J. Cell Biol. 111, 2939‐2949]. We have isolated the cDNA encoding the human PH‐20 protein from a testis library. This cDNA was expressed in RK 13 cells using a vaccinia virus expression system. Cells expressing the human PH‐20 protein possess hyaluronidase activity. Treatment with PI‐PLC releases the hyaluronidase into the the medium with a concomitant large increase in enzymatic activity. These results demonstrate that the human PH‐20 protein has hyaluronidase activity.

Crystal structure of the APC10/DOC1 subunit of the human anaphase-promoting complex.

The anaphase-promoting complex (APC), or cyclosome, is a cell cycle-regulated ubiquitin ligase that controls progression through mitosis and the G1 phase of the cell cycle. The APC is composed of at least 11 subunits; no structure has been determined for any of these subunits. The subunit APC10/DOC1, a one-domain protein consisting of 185 amino acids, has a conserved core (residues 22–161) that is homologous to domains found in several other putative ubiquitin ligases and, therefore, may play a role in ubiquitination reactions. Here we report the crystal structure of human APC10 at 1.6 Å resolution. The core of the protein is formed by a β-sandwich that adopts a jellyroll fold. Unexpectedly, this structure is highly similar to ligand-binding domains of several bacterial and eukaryotic proteins, such as galactose oxidase and coagulation factor Va, raising the possibility that APC10 may function by binding a yet unidentified ligand. We further provide biochemical evidence that the C-terminus of APC10 binds to CDC27/APC3, an APC subunit that contains multiple tetratrico peptide repeats.

Dermal glands of Xenopus laevis contain a polypeptide with a highly repetitive amino acid sequence

Mature dermal glands of Xenopus laevis contain storage granules with a characteristic ellipsoid shape. These granules contain, as a minor component, a heat‐stable, acidic polypeptide with an apparent molecular mass of 75 kDa. Using antibodies against this protein, positive clones were isolated from a cDNA expression library prepared from skin of X. leaevis. One of the cloned cDNAs encodes a pre‐protein with a typical signal sequence and a mature part of 396 amino acids. The protein contains 33 copies of the sequence Gly‐Gly/Glu‐(Ala‐Pro)2–4‐Ala‐Glu. Using the single‐letter code for the four predominant amino acids, we have termed this polypeptide the APEG protein. Near its carboxy‐terminus, one segment has been found with an amino acid sequence similar to that of spasmolytic polypeptide from porcine pancreas and to the human protein pS2.