Application of reverse transcription-PCR to total RNA prepared from TNF-alpha (tumour necrosis factor-alpha)-stimulated HUVECs (human umbilical vein endothelial cells) revealed that the syndecan-2 mRNA was up-regulated by this inflammatory stimulus. By immunoprecipitation using an anti-syndecan-2 antibody on TNF-alpha-stimulated HUVEC lysates, inflammation-induced interleukin-8 was found to be an interaction partner of this HS (heparan sulphate) proteoglycan, but not of any other syndecan on these cells. The glycosylated [Syn2(ect)(+HS)] and non-glycosylated [Syn2(ect)(-HS)] forms of Syn2(ect) (the syndecan-2 ectodomain) were purified from a stably transfected human cell line and from a bacterial expression system respectively. By CD spectroscopy, Syn2(ect) was found to adopt an all-beta secondary structure. The dissociation constant of Syn2(ect)(+HS) with respect to interleukin-8 binding was determined by isothermal fluorescence titrations to be 23 nM. Despite its lack of HS chains, Syn2(ect)(-HS) exhibited significant binding to the chemokine, with a K (d) of >1 microM. Thus, in addition to glycosaminoglycan binding, protein-protein contacts might also contribute to the chemokine-proteoglycan interaction.

The filamentous cyanobacterium Anabaena PCC 7120 (now renamed Nostoc PCC 7120) possesses two genes for superoxide dismutase (SOD). One is an iron-containing (FeSOD) whereas the other is a manganese-containing superoxide dismutase (MnSOD). Localization experiments and analysis of the sequence showed that the FeSOD is cytosolic, whereas the MnSOD is a membranebound homodimeric protein containing one transmembrane helix, a spacer region, and a soluble catalytic domain. It is localized in both cytoplasmic and thylakoid membranes at the same extent with the catalytic domains positioned either in the periplasm or the thylakoid lumen. A phylogenetic analysis revealed that generally the highly homologous MnSODs of filamentous cyanobacteria are unique in being membrane-bound. Two recombinant variants of Anabaena MnSOD lacking either the hydrophobic region (MnSOD(⌬28)) or the hydrophobic and the linker region (MnSOD(⌬60)) are shown to exhibit the characteristic manganese peak at 480 nm, an almost 100% occupancy of manganese per subunit, a specific activity using the ferricytochrome assay of (660 ؎ 90) unit mg ؊1 protein and a dissociation constant for the inhibitor azide of (0.84 ؎ 0.05) mM. Using stopped-flow spectroscopy it is shown that the decay of superoxide in the presence of various (MnSOD(⌬28)) or (MnSOD(⌬60)) concentrations is first-order in enzyme concentration allowing the calculation of catalytic rate constants which increase with decreasing pH: 8 ؋ 10 6

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