TenascinR (TNR) is an extracellular matrix glycoprotein belonging to the tenascin family of adhesion proteins. 1-3 TNR is expressed in the central nervous system by oligodendrocytes and selected inhibitory interneurons. It shows highest expression during the postnatal period of active myelination and promotes neurite outgrowth and synaptic functions. 1, 2 It is essential for formation of perineuronal nets, the mesh-like network of extracellular matrix (ECM) molecules that surrounds some neurons. 4 The 180 kD, 1327 amino acid (aa) form of human TNR contains a signal sequence, three heptad repeats that mediate coiled-coil trimer formation, five EGF-like repeats, nine fibronectin type III repeats (FN), and a C-terminal Ca2+-binding fibrinogen-related domain. TNR isoform 2 (160 kD) lacks a portion of FN#6 (aa # 773-862). 3 Mature human TNR isoform 1 shows 94%, 94%, 93%, 93% and 76% aa identity with cow, mouse, rat, dog and chicken TNR, respectively. Experiments using recombinant TNR fragments indicate that EGF-like domains are counteradhesive for neurons and microglia and contribute to their migration. 1, 5-7 This region interacts with immunoglobulin superfamily molecules including contactin, phosphacan and voltage-gated sodium channel β subunits. However, the fibronectin domains are adhesive for the lectican family of chondroitin sulfate proteoglycans (brevican, aggrican, versican and neurocan; FN 3-5), contactin (FN 2-3) and sodium channel β subunits (FN 6-8). 6-9 These adhesive interactions can compete with each other, but can also contribute to crosslinking of lecticans and contactin with other ECM molecules to form perineuronal nets. 9, 10 Post-translational modification of TNR can differ with time and location. 11 Notably, glycosylation may include GalNAc-4-SO4, O-linked sialylated glycans, "brain-type" neutral N-glycans and the HNK-1 carbohydrate epitope that is thought to be involved in regulation of synaptic plasticity. 11, 12
Source: Human CD33 Signal Peptide (Met 1-Ala 16), Human TenascinR (Glu 34-Phe 1358), HHHHHH; A DNA sequence encoding the mature human TenascinR (Glu 34-Phe 1358; Accession # Q92752) was fused to the signal peptide from human CD33 at the amino terminus and to a polyhistidine tag at the carboxy-terminus. The chimeric protein was expressed in a mouse myeloma cell line, NS0.
Molecular Mass: Based on N-terminal sequencing, the recombinant human TenascinR starts at Glu 34 and has a calculated molecular mass of approximately 146.8 kD. As a result of glycosylation, the recombinant human TenascinR migrates as an approximately 157-180 kD protein in SDS-PAGE under reducing conditions.
Endotoxin Level: < 1.0 EU per 1 μg of the protein as determined by the LAL method.
Activity: Measured by its ability to bind rhContactin-1/Fc in a functional ELISA. rhTenascin-R, immobilized at 5ug/ml (100 μL/well), will bind rhContactin-1/Fc with an apparent KD < 10 nM.
Reconstitution: It is recommended that sterile PBS be added to the vial to prepare a working stock solution of no less than 100ug/ml. The carrier-free protein should be used immediately upon reconstitution to avoid losses in activity due to non-specific binding to the inside surface of the vial. For long term storage as a dilute solution, a carrier protein (e.g. 0.1% HSA or BSA) should be added to the vial.
Storage and Stability: Lyophilized samples are stable for up to twelve months from date of receipt at -20°C. Upon reconstitution, this protein can be stored under sterile conditions at 2°-8° C for one month or at -20°C in a manual defrost freezer for three months without detectable loss of activity. Avoid repeated freeze-thaw cycles.
