AbstractAims To define further the role of IRS‐1 mutations in human syndromes of severe insulin resistance.Methods The IRS‐1 gene was scanned for mutations in 83 unrelated affected subjects and 47 unaffected individuals using fluorescent single‐strand conformation polymorphism (fSSCP) analysis. A novel heterozygous mutation, Gly1158Glu, was found in one affected subject. Four and two subjects were heterozygous for the previously reported variants Gly972Arg and Ala513Pro, respectively. The previously identified variant Gly819Arg was found in one affected and one unaffected subject. While Gly972Arg has been described to alter the signalling properties of IRS‐1, no functional studies of Ala513Pro or Gly1158Glu have been reported.Results Chinese hamster ovary (CHO) cells stably over‐expressing the insulin receptor were transiently transfected with vectors expressing either wild‐type, Glu1158 or Pro513 IRS‐1. A modest increase in insulin‐stimulated tyrosine phosphorylation of Glu1158 IRS‐1 was observed. However, this did not result in any significant change in the association of Grb2 or the p85α subunit of PI3‐kinase or of PI3‐kinase activity. In parallel studies, the Pro513 IRS‐1 variant was indistinguishable from wild‐type IRS‐1.Conclusions While subtle effects of these variants cannot be excluded in this system, it is unlikely that these variants are responsible for the extreme insulin resistance seen in the subjects harbouring them. Although IRS proteins play a central role in insulin signalling, functionally significant mutations in the IRS‐1 gene are a rare cause of human syndromes of severe insulin resistance.