The N-terminal globular domain of the laminin α1 chain binds to α1β1 and α2β1 integrins and to the heparan sulfate-containing domains of perlecan

AbstractThe N-terminal domains VI plus V (62 kDa) and V alone (43 kDa) of the laminin α1 chain were obtained as recombinant products and shown to be folded into a native form by electron microscopy and immunological assays. Domain VI alone, which corresponds to an LN module, did not represent an autonomously folding unit in mammalian cells, however. Fragment α1VI/V, but not fragment α1V, bound to purified α1β1 and α2β1 integrins, to heparin, and to heparan sulfate-substituted domains I and V of perlecan. This localized the binding activities to the LN module, which contains two basic sequences suitable for heparin interactions

Complete sequence, recombinant analysis and binding to laminins and sulphated ligands of the N-terminal domains of laminin alpha3B and alpha5 chains.

The N-terminal sequences of mouse laminin alpha3B and alpha5 chains have been completed and demonstrate the presence of a signal peptide followed by a complete laminin N-terminal (LN) module (domain VI). These signal peptides were released after recombinant production of larger fragments comprising domains VI/V (45-65 kDa) from this region yielding properly folded proteins, which were secreted from HEK-293-EBNA cells. Pepsin digestion of these fragments yielded products of 25-35 kDa, which consisted only of domain V. The alphaVI/V fragments were able to inhibit self-assembly of laminin-1, with those from the alpha3B and alpha5 chains being more active than those from alpha1 and alpha2 chains. Domain V fragments, however, showed a reduced activity, indicating the major contribution of the LN module in inhibition. These interactions were confirmed by surface-plasmon-resonance assays demonstrating moderate affinities (K(d)=0.02 to >6 microM) for the binding to laminin-1. This indicated that laminins containing alpha3B or alpha5 chains should also be able to form non-covalent networks by polymerization. The LN modules also showed heparin binding in affinity chromatography, which was strongest for alpha1/alpha2, moderate for alpha3B, whereas no binding was observed for alpha5. They all bound to heparan sulphate chains of perlecan and to sulphatides, with a lower variability in binding activity. Specific antibodies were raised against alpha3BVI/V and alpha5VI/V and were shown to stain basement membrane zones in various mouse tissues. These antibodies also allowed the identification of a new laminin assembly form 5B consisting of alpha3B, beta3 and gamma2 chains

Limited cleavage of extracellular matrix protein BM-40 by matrix metalloproteinases increases its affinity for collagens

The 33-kDa matrix protein BM-40 (SPARC, osteonectin) consists of an acidic N-terminal domain I, a central cysteine-rich follistatin-like module, and a C-terminal extracellular calcium-binding (EC) module. Previous studies attributed collagen IV and high affinity calcium binding of BM-40 to its EC module, which was shown by x-ray crystallography to consist of an EF-hand pair surrounded by several α-helical and loop segments. This module was now shown by surface plasmon resonance assay to bind with similar affinities to collagens I, III, and V. Cleavage of recombinant BM-40 and its EC module by collagenase-3, gelatinases A and B, matrilysin, and stromelysin-1 showed similar fragment patterns, whereas collagenase-1 was inactive. Some differences were, however, observed in cleavage rates and the preference of certain cleavage sites. Edman degradation of fragments demonstrated only three to four major cleavage sites in the central region of domain I and a single uniform cleavage in helix C of the EC module. Cleavage is accompanied by a 7-20-fold increase in binding activity for collagens I, IV, and V but revealed only small effects on calcium-dependent α-helical changes in the EC module. The data were interpreted to indicate that helix C cleavage is mainly responsible for enhancing collagen affinity by exposing the underlying helix A of the EC module. A similar activation may also occur in situ as indicated previously for tissue-derived BM-4

Cluster categories combining molecular and epidemiological information and cluster level analysis in Baden-Württemberg (Germany).

<p>Cluster categories combining molecular and epidemiological information and cluster level analysis in Baden-Württemberg (Germany).</p

Integration of molecular typing results_pilot Study_Germany

<p>This is the supporting data file for the manuscript entitled "Integration of molecular typing results into tuberculosis surveillance in Germany ─ a pilot study" (Andrés et al., PLoS ONE). The dataset includes anonymized, routinely collected notification data for 918 individuals. The data are published open-access, in accordance with the PLoS ONE data policy (2017).</p

Age distribution of clustered and non-clustered TB cases in Baden-Württemberg (Germany), categorized according to origin.

<p>GB: German-born; FB: foreign-born.</p

Molecular cluster characteristics in Baden-Württemberg (Germany) by A. Geographical distribution and B. Patient origin.

<p>Number of clusters; Size 2 (n = 60); Size 3 (n = 13); Size 4 (n = 6); Size >5 (n = 6).</p

Pilot data flow of the molecular typing results in Baden Württemberg (Germany).

<p>1. Molecular typing requested by local public health offices to NRL; 2. Culture shipment to NRL; 3. Communication of molecular cluster information from NRL to local public health offices and 4. Communication of full molecular typing results from NRL to RKI.</p

Factors associated with TB patients in “No epi links” clusters compared to patients in all other cluster categories in Baden-Württemberg (Germany).

<p>Factors associated with TB patients in “No epi links” clusters compared to patients in all other cluster categories in Baden-Württemberg (Germany).</p