A mutational analysis of receptor binding sites of interleukin-1 beta: differences in binding of human interleukin-1 beta muteins to human and mouse receptors.
The 3-D crystal structure of interleukin-1 beta (IL-1 beta) has been used to define its receptor binding surface by mutational analysis. The surface of IL-1 beta was probed by site-directed mutagenesis. A total of 27 different IL-1 beta muteins were constructed, purified and analyzed. Receptor binding measurements on mouse and human cell lines were performed to identify receptor affinities. IL-1 beta muteins with modified receptor affinity were evaluated for structural integrity by CD spectroscopy or X-ray crystallography. Changes in six surface loops, as well as in the C- and N-termini, yielded muteins with lower binding affinities. Two muteins with intact binding affinities showed 10- to 100-fold reduced biological activity. The surface region involved in receptor binding constitutes a discontinuous area of approximately 1000 A2 formed by discontinuous polypeptide chain stretches. Based on these results, a subdivision into two distinct local areas is proposed. Differences in receptor binding affinities for human and mouse receptors have been observed for some muteins, but not for wild-type IL-1 beta. This is the first time a difference in binding affinity of IL-1 beta muteins to human and mouse receptors has been demonstrated.
Grütter
M G
MG
Department of Biotechnology, Ciba-Geigy Ltd, Basle, Switzerland.
van Oostrum
J
J
Priestle
J P
JP
Edelmann
E
E
Joss
U
U
Feige
U
U
Vosbeck
K
K
Schmitz
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007375:Interleukin-1 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D017472:Receptors, Interleukin-1 / Q000737:chemistry / Q000378:metabolism*; D013329:Structure-Activity Relationship
D007375:Interleukin-1; D017472:Receptors, Interleukin-1