Molecular cloning of the guinea-pig IL-5 receptor alpha and beta subunits and reconstitution of a high affinity receptor.
The functional IL-5 receptor is a heteromeric complex consisting of an alpha and beta subunit. The cloning, sequencing and expression of guinea-pig IL-5Ralpha and beta subunits is described. The guinea-pig IL-5Ralpha subunit cDNA encodes a protein of M(r)47 kDa, which is 72 and 66% homologous to the human and murine orthologs, respectively. Three guinea-pig IL-5Rbeta subunit cDNA clones were isolated, which differ in the N-terminus and are 56-64% homologous to the human and murine IL-5Rbeta subunits. Expressing human IL-5Ralphabeta and guinea-pig IL-5Ralphabeta(1)in the baculovirus-insect cell system resulted in recombinant receptors which bound hIL-5 with high affinity (K(d)=0.19 and 0.11 nM, respectively). Expressing just gpIL-5Ralpha was not sufficient to demonstrate binding. This contrasts with the human receptor, where hIL-5Ralpha alone can bind hIL-5 with high affinity. gpIL-5Ralphabeta(1)bound both hIL-5 and mIL-5 with comparable affinity (K(i)=0.10 and 0.06 nM), similar to that seen with hIL-5Ralphabeta. Thus, both the heteromeric hIL-5R and gpIL-5Ralphabeta(1)can bind multiple IL-5 orthologs with high affinity whereas the murine IL-5R is selective for the murine ligand.
Scott
C W
CW
Respiratory, Inflammation and Neuroscience Department, AstraZeneca Pharmaceuticals, Wilmington, DE 19850, USA.
Logsdon
N J
NJ
Wilkins
D E
DE
Norris
T E
TE
Sobotka-Briner
C
C
Hubbs
S
S
Graham
A
A
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D003001:Cloning, Molecular; D015870:Gene Expression; D006168:Guinea Pigs; D006801:Humans; D015848:Interleukin-5 / Q000378:metabolism*; D008969:Molecular Sequence Data; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D018123:Receptors, Interleukin / Q000235:genetics* / Q000378:metabolism; D053649:Receptors, Interleukin-5; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA; D018411:Spodoptera / Q000166:cytology
D015848:Interleukin-5; D020033:Protein Isoforms; D018123:Receptors, Interleukin; D053649:Receptors, Interleukin-5; D011993:Recombinant Fusion Proteins