| Many tissues such as muscle, skin, liver, and peripheral nerve, have remarkable ability to repair and regrow after injury. However, the CNS (brain and spinal cord) is limited in its ability to repair or regrowth causing permanent brain damage or paralyses. Most recently an inhibitory myelin protein, Nogo (Neurite outgrowth), has been cloned and characterized. It may help block the regeneration of the CNS. Nogo is the 4th member of reticulon (Rtn) family. There are three alternative isoforms of Nogo, designated Nogo-A (full length), an intermediate form Nogo-B (373 aa, ~37 K, lacks 186-1004 aa within the extracellular domain), and a shorter form Nogo-C (199 aa, ~25 K, similar to rat vp20 and foocen-s, lacks 186-1004 aa but which has a smaller, alternative N-terminal domain). A 66-aa hydrophilic region of Nogo (Nogo-66), located between the two TM domains, has the most inhibitory properties of Nogo. In contrast to Nogo, Rtn 1, -2, and 3 do not inhibit axonal regeneration. This 66-aa region also has the least similarity to Rtn proteins. The corresponding Rtn-P4 peptide sequence has no activity. Recently, a brain specific leucine-rich-repeat protein with high affinity for soluble Nogo-66, termed Nogo receptor (Ngr) has been cloned and characterized. Cleavage of Ngr from the axonal surface renders neurons insensitive to Nogo-66. Ngr expression is sufficient to impart Nogo-66 axonal inhibition to unresponsive neurons. Ngr protein (mouse, rat, monkey, and human 473 aa, chromosome 22q11) contains a signal sequence followed by eight LRR domains, an LRR cysteine-rich CT-flanking domain, and unique GPI anchorage site. Human and mouse Ngr proteins are ~89% identical. Ngr is expressed in brain and lower levels are also detected in hear and kidney but not in other peripheral tissues. Ngr is localized to axons. Source: Human Ngr 27-447aa (extracellular domain) was fused with human IgG (100-330aa), expressed as fusion protein in NSO myeloma cells. Recombinant protein chimera is a disulfide-linked homodimer. Non-glycosylated Ngr-Fc monomer calculated wt is ~72kD. Due to glycosylation, Ngr-Fc protein has a mol wt of ~95-100kD under reducing conditions. Applications: Suitable for use in Western Blot. This preparation of Ngr-Fc protein is denatured and not biologically active. It is also not suitable for coating on ELISA plates or applications other than Western. Other applications not tested. Recommended Dilution: Load 5-10ul/lane to visualize with antibodies. Heat once prior to electrophoresis. Optimal dilutions to be determined by the researcher. Storage and Stability: May be stored at 4C for short-term only. For long-term storage, store at -20C. Aliquots are stable for at least 6 months at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer. |
