American Diabetes Association

TABLE 2

Effect of insulin infusion on phosphorylation of endogenous IRS-1 isolated from muscle biopsies

Residues	Sequence	Site(s)	Fold change due to insulin
303–325	(R)SRTESITATpSPASM_oxVGGKPGSFR	Ser³¹²	2.6 ± 0.4^*
328–353	(R)ASpSDGEGTM_oxSRPASVDGSPVSPSTNR	Ser³³⁰	1.1 ± 0.2
328–353	(R)ASSDGEGTM_oxSRPASVDGSPVpSPSTNR	Ser³⁴⁸	0.4 ± 0.1^*
444–457	(R)SVpTPDSLGHTPPAR	Thr⁴⁴⁶	0.2 ± 0.1^*
494–520	(R)CpTPGTGLGTSPALAGDEAASAADLDNR	Thr⁴⁹⁵	0.1 ± 0.1^*
573–580	(R)HpSAFVPTR	Ser⁵⁷⁴	1.1 ± 0.1
596–626	(R)GGHHRPDSSTLHTDDGYM_oxPM_oxpSPGVAPVPSGR	Ser⁶¹⁶	2.9 ± 0.8^*
627–638	(R)KGpSGDYM_oxPM_oxSPK	Ser⁶²⁹	1.2 ± 0.1
627–638	(R)KGSGDYM_oxPM_oxpSPK	Ser⁶³⁶	2.1 ± 0.3^*
892–922	(PK)pSPGEYVNIEFGSDQSGYLSGPVAFHSSPSVR	Ser⁸⁹²	1.6 ± 0.1^*
999–1016	(R)QSYVDTpSPAAPVSYADM_oxR	Ser¹⁰⁰⁵	0.3 ± 0.2^*
1075–1081	(R)VNLpSPNR	Ser¹⁰⁷⁸	1.1 ± 0.1
1098–1112	(R)RHpSSETFSSTPSATR	Ser¹¹⁰⁰	0.8 ± 0.2
1099–1112	(R)HSpSETFSSTPSATR	Ser¹¹⁰¹	1.3 ± 0.1^*
1221–1236	(R)RSpSEDLSAYASISFQK	Ser¹²²³	1.3 ± 0.1^*
303–325	(R)SRTEpSITATSPASM_oxVGGKPGpSFR	Ser³⁰⁷	NA^†
303–325	(R)SRTESITATSPASM_oxVGGKPGpSFR	Ser³²³	NA^†
525–538	(R)THpSAGTSPTITHQK	Ser⁵²⁷	NA^†
525–538	(R)THSAGTpSPTITHQK	Ser⁵³¹	NA^†
1141–1161	(R)HpSSASFENVWLRPGELGGAPK	Ser¹¹⁴²	NA^†
1141–1161	(R)HSpSASFENVWLRPGELGGAPK	Ser¹¹⁴³	NA^†
1141–1161	(R)HSSApSFENVWLRPGELGGAPK	Ser¹¹⁴⁵	NA^†

Residues	Sequence	Site(s)	Fold change due to insulin
303–325	(R)SRTESITATpSPASM_oxVGGKPGSFR	Ser³¹²	2.6 ± 0.4^*
328–353	(R)ASpSDGEGTM_oxSRPASVDGSPVSPSTNR	Ser³³⁰	1.1 ± 0.2
328–353	(R)ASSDGEGTM_oxSRPASVDGSPVpSPSTNR	Ser³⁴⁸	0.4 ± 0.1^*
444–457	(R)SVpTPDSLGHTPPAR	Thr⁴⁴⁶	0.2 ± 0.1^*
494–520	(R)CpTPGTGLGTSPALAGDEAASAADLDNR	Thr⁴⁹⁵	0.1 ± 0.1^*
573–580	(R)HpSAFVPTR	Ser⁵⁷⁴	1.1 ± 0.1
596–626	(R)GGHHRPDSSTLHTDDGYM_oxPM_oxpSPGVAPVPSGR	Ser⁶¹⁶	2.9 ± 0.8^*
627–638	(R)KGpSGDYM_oxPM_oxSPK	Ser⁶²⁹	1.2 ± 0.1
627–638	(R)KGSGDYM_oxPM_oxpSPK	Ser⁶³⁶	2.1 ± 0.3^*
892–922	(PK)pSPGEYVNIEFGSDQSGYLSGPVAFHSSPSVR	Ser⁸⁹²	1.6 ± 0.1^*
999–1016	(R)QSYVDTpSPAAPVSYADM_oxR	Ser¹⁰⁰⁵	0.3 ± 0.2^*
1075–1081	(R)VNLpSPNR	Ser¹⁰⁷⁸	1.1 ± 0.1
1098–1112	(R)RHpSSETFSSTPSATR	Ser¹¹⁰⁰	0.8 ± 0.2
1099–1112	(R)HSpSETFSSTPSATR	Ser¹¹⁰¹	1.3 ± 0.1^*
1221–1236	(R)RSpSEDLSAYASISFQK	Ser¹²²³	1.3 ± 0.1^*
303–325	(R)SRTEpSITATSPASM_oxVGGKPGpSFR	Ser³⁰⁷	NA^†
303–325	(R)SRTESITATSPASM_oxVGGKPGpSFR	Ser³²³	NA^†
525–538	(R)THpSAGTSPTITHQK	Ser⁵²⁷	NA^†
525–538	(R)THSAGTpSPTITHQK	Ser⁵³¹	NA^†
1141–1161	(R)HpSSASFENVWLRPGELGGAPK	Ser¹¹⁴²	NA^†
1141–1161	(R)HSpSASFENVWLRPGELGGAPK	Ser¹¹⁴³	NA^†
1141–1161	(R)HSSApSFENVWLRPGELGGAPK	Ser¹¹⁴⁵	NA^†

Data are shown as fold change compared with basal (before insulin infusion values; see text for calculations) and are presented as means ± SEM of four independent experiments. Each experiment represents the average of two replicates from the same muscle homogenate.

P < 0.05 by paired t test (basal vs. insulin, comparing normalized ion intensities).

†

Monophosphorylated peptides were not sufficiently separated by HPLC to permit individual quantification (see text). Evidence for the existence of modification at each site was obtained by tandem mass spectrometry. M_ox, oxidized methionine; pS, phosphoserine; pT, phosphothreonine.

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