title: Cysteine-specific Cu2+ chelating tags used as paramagnetic probes in double electron electron resonance.
abstract: Double electron electron resonance (DEER) is an attractive technique that is utilized for gaining insight into protein structure and dynamics via nanometer-scale distance measurements. The most commonly used paramagnetic tag in these measurements is a nitroxide spin label, R1. Here, we present the application of two types of high-affinity Cu(2+) chelating tags, based on the EDTA and cyclen metal-binding motifs as alternative X-band DEER probes, using the B1 immunoglobulin-binding domain of protein G (GB1) as a model system. Both types of tags have been incorporated into a variety of protein secondary structure environments and exhibit high spectral sensitivity. In particular, the cyclen-based tag displays distance distributions with comparable distribution widths and most probable distances within 1-3 Å when compared to homologous R1 distributions. The results display the viability of the cyclen tag as an alternative to the R1 side chain for X-band DEER distance measurements in proteins.
authors.affiliation: Department of Chemistry, University of Pittsburgh , 219 Parkman Avenue, Pittsburgh, Pennsylvania 15260, United States.
authors.forename: Timothy F
authors.initials: TF
authors.lastname: Cunningham
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Shannon
authors.forename: Miriam R
authors.initials: MR
authors.lastname: Putterman
authors.forename: Rajith J
authors.initials: RJ
authors.lastname: Arachchige
authors.forename: Ishita
authors.initials: I
authors.lastname: Sengupta
authors.forename: Min
authors.initials: M
authors.lastname: Gao
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Jaroniec
authors.forename: Sunil
authors.initials: S
authors.lastname: Saxena
mesh_terms: D002412:Cations; D002614:Chelating Agents; D003300:Copper; D000080027:Cyclams; D003545:Cysteine; D004492:Edetic Acid; D006571:Heterocyclic Compounds; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry; D013113:Spin Labels
chemical_list: D002412:Cations; D002614:Chelating Agents; D000080027:Cyclams; D006571:Heterocyclic Compounds; D011506:Proteins; D013113:Spin Labels; D003300:Copper; C038072:cyclen; D004492:Edetic Acid; D003545:Cysteine