PMID- 8654370
OWN - NLM
STAT- MEDLINE
DCOM- 19960730
LR  - 20181113
IS  - 0261-4189 (Print)
IS  - 1460-2075 (Electronic)
IS  - 0261-4189 (Linking)
VI  - 15
IP  - 11
DP  - 1996 Jun 3
TI  - Engineering human interleukin-6 to obtain variants with strongly enhanced 
      bioactivity.
PG  - 2726-37
AB  - Interleukin-6 (IL-6) triggers the formation of a high affinity receptor complex 
      with the ligand binding subunit IL-6Ralpha and the signal transducing chain 
      gp130. Since the intracytoplasmic region of the IL-6Ralpha does not contribute to 
      signaling, soluble forms of the extracytoplasmic domain (sIL-6Ralpha), potentiate 
      IL-6 bioactivity and induce a cytokine-responsive status in cells expressing 
      gp130 only. This observation, together with the detection of high levels of 
      circulating soluble human IL-6Ralpha (shIL-6Ralpha) in sera, suggests that the 
      hIL-6-shIL-6Ralpha complex is an alternative form of the cytokine. Here we 
      describe the generation of human IL-6 (hIL-6) variants with strongly enhanced 
      shIL-6Ralpha binding activity and bioactivity. Homology modeling and 
      site-directed mutagenesis of hIL-6 suggested that the binding interface for 
      hIL-6Ralpha is constituted by the C-terminal portion of the D-helix and residues 
      contained in the AB loop. Four libraries of hIL-6 mutants were generated by each 
      time fully randomizing four different amino acids in the predicted AB loop. These 
      libraries were displayed monovalently on filamentous phage surface and sorted 
      separately for binding to immobilized shIL-6Ralpha. Mutants were selected which, 
      when expressed as soluble proteins, showed a 10- to 40-fold improvement in 
      shIL-6Ralpha binding; a further increase (up to 70-fold) was achieved by 
      combining variants isolated from different libraries. Interestingly, high 
      affinity hIL-6 variants show strongly enhanced bioactivity on cells expressing 
      gp13O in the presence of shIL-6Ralpha at concentrations similar to those normally 
      found in human sera.
FAU - Toniatti, C
AU  - Toniatti C
AD  - Department of Genetics, Istituto di Ricerche di Biologia Moleculare, Rome, Italy.
FAU - Cabibbo, A
AU  - Cabibbo A
FAU - Sporena, E
AU  - Sporena E
FAU - Salvati, A L
AU  - Salvati AL
FAU - Cerretani, M
AU  - Cerretani M
FAU - Serafini, S
AU  - Serafini S
FAU - Lahm, A
AU  - Lahm A
FAU - Cortese, R
AU  - Cortese R
FAU - Ciliberto, G
AU  - Ciliberto G
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - England
TA  - EMBO J
JT  - The EMBO journal
JID - 8208664
RN  - 0 (Antigens, CD)
RN  - 0 (DNA Primers)
RN  - 0 (IL6ST protein, human)
RN  - 0 (Interleukin-6)
RN  - 0 (Membrane Glycoproteins)
RN  - 0 (Receptors, Interleukin)
RN  - 0 (Receptors, Interleukin-6)
RN  - 133483-10-0 (Cytokine Receptor gp130)
SB  - IM
MH  - Antigens, CD/metabolism
MH  - Bacteriophage M13/genetics
MH  - Base Sequence
MH  - Cytokine Receptor gp130
MH  - DNA Primers/chemistry
MH  - Gene Library
MH  - Genetic Engineering
MH  - Genetic Vectors
MH  - Humans
MH  - Interleukin-6/*chemistry/genetics
MH  - Membrane Glycoproteins/metabolism
MH  - Models, Molecular
MH  - Molecular Sequence Data
MH  - Protein Binding
MH  - Protein Structure, Tertiary
MH  - Receptors, Interleukin/metabolism
MH  - Receptors, Interleukin-6
MH  - Structure-Activity Relationship
PMC - PMC450208
EDAT- 1996/06/03 00:00
MHDA- 1996/06/03 00:01
PMCR- 1997/06/03
CRDT- 1996/06/03 00:00
PHST- 1996/06/03 00:00 [pubmed]
PHST- 1996/06/03 00:01 [medline]
PHST- 1996/06/03 00:00 [entrez]
PHST- 1997/06/03 00:00 [pmc-release]
PST - ppublish
SO  - EMBO J. 1996 Jun 3;15(11):2726-37.