A simple strategy for ¹³C, ¹H labeling at the Ile-γ2 methyl position in highly deuterated proteins.
A straightforward approach for the production of highly deuterated proteins labeled with ¹³C and ¹H at Ile-γ2 methyl positions is described. The utility of the methodology is illustrated with an application involving the half proteasome (360 kDa). High quality 2D Ile ¹³C(γ)²,¹H(γ)² HMQC data sets, exploiting the methyl-TROSY principle, are recorded with excellent sensitivity and resolution, that compare favorably with Ile ¹³C(δ)¹,¹H(δ)¹ spectra. This labeling scheme adds to a growing list of different approaches that are significantly impacting the utility of solution NMR spectroscopy in studies of supra-molecular systems.
Department of Molecular Genetics, The University of Toronto, ON, Canada.
Amy M
AM
Ruschak
Algirdas
A
Velyvis
Lewis E
LE
Kay
D002247:Carbon Isotopes; D003903:Deuterium; D007532:Isoleucine / Q000737:chemistry*; D007553:Isotope Labeling / Q000379:methods*; D007930:Leucine / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D046988:Proteasome Endopeptidase Complex / Q000032:analysis / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D014633:Valine / Q000737:chemistry; D014805:Vitamin B 12 / Q000031:analogs & derivatives / Q000737:chemistry
D002247:Carbon Isotopes; D011506:Proteins; D007532:Isoleucine; C051329:factor III, vitamin B 12; D003903:Deuterium; D046988:Proteasome Endopeptidase Complex; D007930:Leucine; D014633:Valine; D014805:Vitamin B 12