Lysozymes were purified from three invertebrates: a marine bivalve, a marine conch, and an earthworm. The purified lysozymes all showed a similar molecular weight of 13 kDa on SDS/PAGE. Their N-terminal sequences up to the 33rd residue determined here were apparently homologous among them; in addition, they had a homology with a partial sequence of a starfish lysozyme which had been reported before. The complete sequence of the bivalve lysozyme was determined by peptide mapping and subsequent sequence analysis. This was composed of 123 amino acids including as many as 14 cysteine residues and did not show a clear homology with the known types of lysozymes. However, the homology search of this protein on the protein or nucleic acid database revealed two homologous proteins. One of them was a gene product, CELF22 A3.6 of C. elegans, which was a functionally unknown protein. The other was an isopeptidase of a medicinal leech, named destabilase. Thus, a new type of lysozyme found in at least four species across the three classes of the invertebrates demonstrates a novel class of protein/lysozyme family in invertebrates. The bivalve lysozyme, first characterized here, showed extremely high protein stability and hen lysozyme-like enzymatic features.Keywords: lysozyme; invertebrate; purification; characterization; sequence.Lysozyme is a ubiquitous enzyme that is wildly distributed in the animal kingdom [1]. The biological function of this enzyme is believed to be self-defense from bacterial infection because it induces bacterial cell lysis by hydrolyzing b-1,4 linked glycosidic bond of the peptidoglycan on the bacterial cell wall [2].It is known that several types of lysozymes are distributed in the different parts of the animal kingdom. Goose-type and phage-type lysozymes are found in a part of avian [1] and bacterial phages [3], respectively. Other types of lysozymes are also seen in plants [4] and bacteria [5]. Chicken-type lysozyme is the most conventional one, being distributed in a large number of vertebrates [1] from fish to mammals and also found in insects like moths and flies [6]. Jolles and Jolles purified and partially sequenced another invertebrate lysozyme from a marine invertebrate, a starfish called Asterias rubens [7]. However, its N-terminal sequence was not similar to that of any known type of lysozyme including the chicken type lysozyme, implicating a novel type of lysozyme. Other invertebrate lysozymes had been purified and partially characterized [8±11], but the sequence data were not complete.To investigate the structure and function of invertebrate lysozyme, we purified lysozymes from three species of invertebrates: a marine bivalve, a marine conch, and an earthworm. We determined the N-terminal sequences of the three species and the complete sequence of the marine bivalve lysozyme. The biological function and evolutionary importance of this enzyme were discussed on the basis of the sequence and enzymatic properties. EXPERIMENTAL PROCEDURES Purification of bivalve lysozymeMarine bivalves (Tapes ...

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BACKGROUND: The hypothesis that malignant tumours are generated by rare populations of cancer stem cells that are more tumourigenic than other cancer cells has gained increasing credence. The objective of this study was to identify and characterise a subpopulation of human sarcoma-initiating cells. METHODS: We examined established rhabdomyosarcoma cell lines by flow cytometry. Tumourigenesis was examined by xenograft models. Real-time PCR and immunohistochemistry were performed to examine the gene expression using cell lines and biopsy specimens. RESULTS: Rhabdomyosarcoma cell lines included small populations of fibroblast growth factor receptor 3 (FGFR3)-positive cells. FGFR3-positive KYM-1 and RD cells were more strongly tumourigenic than FGFR3-negative cells. In addition, xenoengraftment of 33% of single FGFR3-positive KYM-1 cells yielded tumour formation. Stem cell properties of FGFR3-positive cells were further established by real-time PCR, which demonstrated upregulation of undifferentiated cell markers and downregulation of differentiation markers. We showed that in the absence of serum, addition of basic fibroblast growth factor maintained and enriched FGFR3-positive cells. On the other hand, ciliary neurotrophic factor reduced the proportion of FGFR3-positive cells. Real-time PCR and immunohistochemical examination revealed that embryonal rhabdomyosarcoma patient biopsy specimens were found to overexpress FGFR3. CONCLUSIONS: Our findings suggest that rhabdomyosarcoma cell lines include a minor subpopulation of FGFR3-positive sarcomainitiating cells, which can be maintained indefinitely in culture and which is crucial for their malignancy.

Molecular cloning experiments indicated that BW10KD as a BW allergen was a member of the 2S-albumin multigene family.

The structure and the dissociation reaction of oligomers PrP oligo from reduced human prion huPrP C (23-231) have been studied by 1 H-NMR and tryptophan fluorescence spectroscopy at varying pressure, along with circular dichroism and atomic force microscopy. The 1 H-NMR and fluorescence spectral feature of the oligomer is consistent with the notion that the N-terminal residues including all seven Trp residues, are free and mobile, while residues 105~210, comprising the AGAAAAGA motif and S1-Loop-HelixA-Loop-S2-Loop-HelixC, are engaged in intra-and/ or inter-molecular interactions. By increasing pressure to 200 MPa, the oligomers tend to dissociate into monomers which may be identified with PrP C* , a rare metastable form of PrP C stabilized at high pressure (Kachel et al., BMC Struct Biol 6:16). The results strongly suggest that the oligomeric form PrP oligo is in dynamic equilibrium with the monomeric forms via PrP C* , namely huPrP C  huPrP C*  huPrP oligo .