Fractional 13C enrichment of isolated carbons using [1-13C]- or [2- 13C]-glucose facilitates the accurate measurement of dynamics at backbone Calpha and side-chain methyl positions in proteins

TY - JOUR

T1 - Fractional 13C enrichment of isolated carbons using [1-13C]- or [2- 13C]-glucose facilitates the accurate measurement of dynamics at backbone Calpha and side-chain methyl positions in proteins

AU - Lundström, Patrik

AU - Teilum, Kaare

AU - Carstensen, Tommy

AU - Bezsonova, Irina

AU - Wiesner, Silke

AU - Hansen, D Flemming

AU - Religa, Tomasz L

AU - Akke, Mikael

AU - Kay, Lewis E

N1 - Keywords: Amino Acid Sequence; Amino Acids; Animals; Carbon; Carbon Isotopes; Cattle; Glucose; Humans; Methane; Nuclear Magnetic Resonance, Biomolecular; Proteins

PY - 2007

Y1 - 2007

N2 - A simple labeling approach is presented based on protein expression in [1-(13)C]- or [2-(13)C]-glucose containing media that produces molecules enriched at methyl carbon positions or backbone C(alpha) sites, respectively. All of the methyl groups, with the exception of Thr and Ile(delta1) are produced with isolated (13)C spins (i.e., no (13)C-(13)C one bond couplings), facilitating studies of dynamics through the use of spin-spin relaxation experiments without artifacts introduced by evolution due to large homonuclear scalar couplings. Carbon-alpha sites are labeled without concomitant labeling at C(beta) positions for 17 of the common 20 amino acids and there are no cases for which (13)C(alpha)-(13)CO spin pairs are observed. A large number of probes are thus available for the study of protein dynamics with the results obtained complimenting those from more traditional backbone (15)N studies. The utility of the labeling is established by recording (13)C R (1rho) and CPMG-based experiments on a number of different protein systems.

AB - A simple labeling approach is presented based on protein expression in [1-(13)C]- or [2-(13)C]-glucose containing media that produces molecules enriched at methyl carbon positions or backbone C(alpha) sites, respectively. All of the methyl groups, with the exception of Thr and Ile(delta1) are produced with isolated (13)C spins (i.e., no (13)C-(13)C one bond couplings), facilitating studies of dynamics through the use of spin-spin relaxation experiments without artifacts introduced by evolution due to large homonuclear scalar couplings. Carbon-alpha sites are labeled without concomitant labeling at C(beta) positions for 17 of the common 20 amino acids and there are no cases for which (13)C(alpha)-(13)CO spin pairs are observed. A large number of probes are thus available for the study of protein dynamics with the results obtained complimenting those from more traditional backbone (15)N studies. The utility of the labeling is established by recording (13)C R (1rho) and CPMG-based experiments on a number of different protein systems.

U2 - 10.1007/s10858-007-9158-6

DO - 10.1007/s10858-007-9158-6

M3 - Journal article

C2 - 17554498

SN - 0925-2738

VL - 38

SP - 199

EP - 212

JO - Journal of Biomolecular N M R

JF - Journal of Biomolecular N M R

IS - 3

ER -