Structure of a fatty-acid-binding protein from Bacillus subtilis determined by sulfur-SAD phasing using in-house chromium radiation.
Sulfur single-wavelength anomalous dispersion (S-SAD) and halide-soaking methods are increasingly being used for ab initio phasing. With the introduction of in-house Cr X-ray sources, these methods benefit from the enhanced anomalous scattering of S and halide atoms, respectively. Here, these methods were combined to determine the crystal structure of BsDegV, a DegV protein-family member from Bacillus subtilis. The protein was cocrystallized with bromide and low-redundancy data were collected to 2.5 A resolution using Cr Kalpha radiation. 17 heavy-atom sites (ten sulfurs and seven bromides) were located using standard methods. The anomalous scattering of some of the BsDegV S atoms and Br atoms was weak, thus neither sulfurs nor bromides could be used alone for structure determination using the collected data. When all 17 heavy-atom sites were used for SAD phasing, an easily interpretable electron-density map was obtained after density modification. The model of BsDegV was built automatically and a palmitate was found tightly bound in the active site. Sequence alignment and comparisons with other known DegV structures provided further insight into the specificity of fatty-acid selection and recognition within this protein family.
Nan
Jie
J
National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing, People's Republic of China.
Zhou
Yanfeng
Y
Yang
Cheng
C
Brostromer
Erik
E
Kristensen
Ole
O
Su
Xiao Dong
XD
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D001665:Binding Sites; D001965:Bromides; D002857:Chromium; D018360:Crystallography, X-Ray / Q000379:methods*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010168:Palmitates / Q000378:metabolism; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013455:Sulfur
D001426:Bacterial Proteins; D001965:Bromides; D010168:Palmitates; D011993:Recombinant Fusion Proteins; D002857:Chromium; D013455:Sulfur