Human Neuropilin-1 /NRP1 (glycosylated, InCs) recombinant Protein
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Neuropilin receptor-1, Vascular endothelial growth factor 165 receptor, CD304, VEGF165R, NRP
Description of Human Neuropilin-1 /NRP1 (glycosylated, InCs)
Recombinant human Neuropilin-1 (NRP-1, CD304) produced in insect cells is a glycosylated 82.6 kDa protein with 631aa. It is a 130-140 kDa type I transmembrane glycoprotein that regulates axon guidance and angiogenesis. The human NRP-1 contains a 623 aa extracellular domain (ECD) that shows 92-95% aa identity with mouse, rat, bovine and canine NRP-1. The ECD contains two N-terminal CUB domains (termed a1a2), two domains with homology to coagulation factors V and VIII (b1b2) and a MAM (meprin) domain. C-terminally divergent splice variants with 704, 644, 609, and 551 aa lack the MAM and TM domains and are demonstrated or presumed to be soluble antagonists. Heparin, the heparin-binding forms of VEGF (VEGF165, VEGF-B; VEGF-E), PlGF-2, and the C-terminus of Sema3 bind the b1b2 region. NRP-1 and NRP-2 share 48% aa identity within the ECD and can form homo and hetero-oligomers via interaction of their MAM domains. Neuropilins show partially overlapping expression in neuronal and endothelial cells during development. Both neuropilins act as coreceptors with Plexins, mainly Plexin A3 and A4, to bind class III Semaphorins that mediate axon repulsion. However, only NRP-1 binds Sema3A, and only NRP-2 binds Sema 3F. Both are co-receptors with VEGFR-2 (KDR7Flk1) for VEGF165 binding. Sema 3A signaling can be blocked by VEGF165, which has higher affinity for NRP-1. NRP-1 is preferentially expressed in arteries during development or those undergoing remodeling. NRP-1 is also expressed on dendritic cells and mediates DC-induced T-cell proliferation. Recently Neuropilin-1 has been identified as a factor that can facilitate SARS-COV-2 entry into the cells’ interior.
The biological activity was measured by its binding ability in a functional ELISA. Immobilized soluble Neuropilin-1 binds all VEGF-A isoforms except for VEGF121 (Fig 2, see below).
We recommend a quick spin followed by reconstitution in sterile-filtered water or PBS to a concentration of at least 100 µg/ml. Do not vortex.
The lyophilized protein is stable for greater than six months at -20° C to -70° C. After the first thawing we recommend to aliquot the material, because repeated freeze-thaw cycles will decrease the activity. The reconstituted protein should not be stored at 4°C for longer than 2 days. Please avoid repeated freeze-thaw cycles.
Amino Acid Sequence
FRNDKCGDTI KIESPGYLTS PGYPHSYHPS EKCEWLIQAP DPYQRIMINF NPHFDLEDRD CKYDYVEVFD GENENGHFRG KFCGKIAPPP VVSSGPFLFI KFVSDYETHG AGFSIRYEIF KRGPECSQNY TTPSGVIKSP GFPEKYPNSL ECTYIVFAPK MSEIILEFES FDLEPDSNPP GGMFCRYDRL EIWDGFPDVG PHIGRYCGQK TPGRIRSSSG ILSMVFYTDS AIAKEGFSAN YSVLQSSVSE DFKCMEALGM ESGEIHSDQI TASSQYSTNW SAERSRLNYP ENGWTPGEDS YREWIQVDLG LLRFVTAVGT QGAISKETKK KYYVKTYKID VSSNGEDWIT IKEGNKPVLF QGNTNPTDVV VAVFPKPLIT RFVRIKPATW ETGISMRFEV YGCKITDYPC SGMLGMVSGL ISDSQITSSN QGDRNWMPEN IRLVTSRSGW ALPPAPHSYI NEWLQIDLGE EKIVRGIIIQ GGKHRENKVF MRKFKIGYSN NGSDWKMIMD DSKRKAKSFE GNNNYDTPEL RTFPALSTRF IRIYPERATH GGLGLRMELL GCEVEAPTAG PTTPNGNLVD ECDDDQANCH SGTGDDFQLT GGTTVLATEK PTVIDSTIQS GIKLEHHHHH H
Fig. 1.: SDS-PAGE analysis of recombinant human soluble NRP-1. Sample was loaded in 10% SDS-polyacrylamide gel under reducing conditions and stained with Coomassie blue.
Fig. 2: Binding of VEGF-A isoforms to recombinant human soluble Neuropilin-1 receptor (NRP-1) in a functional ELISA. NRP-1 was coated with 1µg/ml (100µl/well), the ligands were added with 10ng/ml.
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