Besides the mnnomcric mammalian 95 kDa progelatinase, two additional forms, ~1 disulfide-bridged 220 kDa dinxr and a 125 kDn form were isolated from human PMN Icukocytes. The I25 kDa progelatinase was identified as a covalcntly linked, disul!!de-bridged hetcrodimer formed of the monomer with a 2s kDa protein. This 25 kDa protein was isolntcd from pclatinase bound to the atIinity support of gelatin-Scpharosc and eluted by DTE-containing bulli3r. The amino acid sequence of tryptic peptides of this protein rcvenled homology with an c+-microglobulin-related protein from rats, a protein so far unknown in humans.

Synaptophysin is a major glycoprotein of Mr approximately 38,000 (in deglycosylated form: Mr approximately 34,000) characteristic of a certain class of small (30‐80 nm diameter) neurosecretory vesicles, including presynaptic vesicles, but also vesicles of various neuroendocrine cells of both neuronal and epithelial phenotype. Using synaptophysin‐specific antibodies we have isolated cDNA clones from rat nervous tissue libraries, which identify an approximately 2.5‐kb mRNA in rat and human cells, including neuroendocrine tumours, that contains a reading frame for a polypeptide of 307 amino acids with a total mol. wt of 33 312. The deduced amino acid sequence, which was partly confirmed by comparison with sequences of two tryptic peptides obtained from purified synaptophysin, revealed four hydrophobic regions of 24 amino acids each, which are characterized, according to conformation prediction analyses, by marked alpha‐helicity. The sequence shows a single potential N‐glycosylation site, which is assigned to the vesicle interior, and a carboxy‐terminal tail of 89 amino acids which contains glycine‐rich tetrapeptide repeats, the epitope of monoclonal antibody SY38, and a number of collagenase‐sensitive sites accessible on the surface of the intact vesicles. These features suggest that the polypeptide spans the vesicle membrane four times, with both N and C termini located on the outer, i.e. cytoplasmic, surface of the vesicles.

Acid sphingomyelinase (sphingomyelin phosphodiesterase, EC 3.1.4.12) was purified from human urine and 12 tryptic peptides were microsequenced (128 residues). Based on regions of minimal codon redundancy, four oligonucleotide mixtures were synthesized and oligonucleotide mixture 1 (20mer; 256 mix) was used to screen 3 X 10(6) independent recombinants from a human fibroblast cDNA library. Putative positive clones (92) were purified and analyzed by Southern hybridization with oligonucleotide mixtures 2‐4. These studies revealed two groups of clones; group 1 (80 clones; inserts ranging from approximately 1.2 to 1.6 kb) hybridized with oligonucleotides mixtures 1‐4, while group II (12 clones; inserts ranging from approximately 1.2 to 1.4 kb) hybridized with oligonucleotide mixtures 1‐3. Several group II clones had larger inserts than those in group I, but did not hybridize with oligonucleotide mixture 4. Screening of a human placental cDNA library with a 450 bp group I fragment, also resulted in the isolation of group I and II clones. Representative clones from group I (pASM‐1) and group II (pASM‐2) were sequenced. pASM‐1 contained a 1879 bp insert which was colinear with 96 microsequenced amino acids, while the pASM‐2 1382 bp insert was colinear with 78 microsequenced residues. Notably, pASM‐2 did not have an internal 172 bp sequence encoding 57 amino acid residues, but had instead an in‐frame 40 bp sequence encoding 13 amino acids which was not present in pASM‐1. These findings demonstrate the presence of two distinct acid sphingomyelinase transcripts in human fibroblasts and placenta and suggest the occurrence of alternative processing of the mRNA encoding this lysosomal hydrolase.

We present the nucleotide sequences and derived amino acid sequences of cDNAs that encode the complete precursors of the extrinsic '23 kDa' and '16 kDa' polypeptides associated with the photosynthetic oxygenevolving complex from spinach. The luminal proteins consist of 267/186 (precursor/mature 23 kDa protein) and 232/149 (16 kDa polypeptide) amino acid residues corresponding to molecular masses of 28.5/20.2 and 24.9/16.5 kDa, respectively. Secondary structure predictions disclose epitopes that are potential candidates for two-step processing of the precursors during import and intraorganelle routing as well as for calcium sequestering, chloride binding and subunit/subunit interaction.Photosynthesis; Oxygen evolution; 23 kDa protein; 16 kDa protein; cDNA nucleotide sequence; Transit peptide; (Spinach)

No abstract