Structure-function studies of interleukin 15 using site-specific mutagenesis, polyethylene glycol conjugation, and homology modeling

J Biol Chem. 1997 Jan 24;272(4):2312-8. doi: 10.1074/jbc.272.4.2312.


Interleukin (IL)-15 is a multifunctional cytokine that shares many biological activities with IL-2. This functional overlap, as well as receptor binding subunits shared by IL-15 and IL-2, suggests tertiary structural similarities between these two cytokines. In this study, recombinant human IL-15 was PEGylated via lysine-specific conjugation chemistry in order to extend the circulation half-life of this cytokine. Although PEGylation did extend the beta-elimination circulation half-life of IL-15 by greater than 50-fold, the biological activity of polyethylene glycol (PEG)-IL-15 was significantly altered. Specifically, PEG-IL-15 lost its ability to stimulate the proliferation of CTLL but took on the properties of a specific IL-15 antagonist in vitro. In comparing sequence alignments and molecular models for IL-2 and IL-15, it was noted that lysine residues resided in regions of IL-15 that may have selectively disrupted receptor subunit binding. We hypothesized that PEGylation of IL-15 interferes with beta but not alpha receptor subunit binding, resulting in the IL-15 antagonist activity observed in vitro. The validity of this hypothesis was tested by engineering site-specific mutants of human IL-15 as suggested by the IL-15 model (IL-15D8S and IL-15Q108S block beta and gamma receptor subunit binding, respectively). As with PEG-IL-15, these mutants were unable to stimulate CTLL proliferation but were able to specifically inhibit the proliferation of CTLL in response to unmodified IL-15. These results supported our model of IL-15 and confirmed that interference of beta receptor subunit binding by adjacent PEGylation could be responsible for the altered biological activity observed for PEG-IL-15.

MeSH terms

  • Amino Acid Sequence
  • Electrophoresis, Polyacrylamide Gel
  • Humans
  • Interleukin-15 / chemistry*
  • Interleukin-15 / metabolism
  • Interleukin-15 / pharmacokinetics
  • Interleukin-2 / chemistry
  • Models, Chemical*
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Polyethylene Glycols / metabolism*
  • Protein Conformation
  • Sequence Alignment
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Structure-Activity Relationship


  • Interleukin-15
  • Interleukin-2
  • Polyethylene Glycols