Our staff maintains and operates a series of sophisticated biophysical instrumentation. Among the analytical methods available to intramural researchers are: analytical ultracentrifugation, isothermal titration calorimetry, surface plasmon resonance, differential scanning calorimetry, dynamic light scattering, circular dichroism spectroscopy, and asymmetric flow field-flow fractionation. In order to gain insight into the formation of signalling clusters in T-cell activation, our resource, in collaboration with NCI and the NIBIB, has applied analytical ultracentrifugation (AUC) and isothermal titration calorimetry (ITC) toward characterizing the binding of the SH2 domain of SLP76 to specific peptide fragments of ADAP having controlled ratios of phosphorylation at 2 distant binding-site tyrosine residues. In collaboration with the NICHD, analytical ultracentrifugation sedimentation velocity has been used to quantify the strength of high-affinity (low nanomolar Kd) self-association for the amino terminal domain (ATD) of AMPA subtype glutamate receptor ion channels, major mediators of fast excitatory synaptic transmission in the human brain. In collaboration with two different labs within the NIAID, we have utilized sedimentation equilibrium and sedimentation velocity approaches of AUC to investigate the association state of various secondary amyloidosis protein (SAA) constructs, and to probe the assembly state of malaria merozoite surface protein 3 (MSP3).

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National Institute of Biomedical Imaging and Bioengineering
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Jensen, Jaime L; Balbo, Andrea; Neau, David B et al. (2015) Mechanistic Implications of the Unique Structural Features and Dimerization of the Cytoplasmic Domain of the Pseudomonas Sigma Regulator, PupR. Biochemistry 54:5867-77
Zhao, Huaying; Balbo, Andrea; Metger, Howard et al. (2014) Improved measurement of the rotor temperature in analytical ultracentrifugation. Anal Biochem 451:69-75
May, Nathan A; Wang, Qiuhong; Balbo, Andrea et al. (2014) Human herpesvirus 7 U21 tetramerizes to associate with class I major histocompatibility complex molecules. J Virol 88:3298-308
Ghirlando, Rodolfo; Zhao, Huaying; Balbo, Andrea et al. (2014) Measurement of the temperature of the resting rotor in analytical ultracentrifugation. Anal Biochem 458:37-9
Zhao, Huaying; Berger, Anthony J; Brown, Patrick H et al. (2013) Analysis of high-affinity assembly for AMPA receptor amino-terminal domains. J Gen Physiol 141:747-9
Coussens, Nathan P; Hayashi, Ryo; Brown, Patrick H et al. (2013) Multipoint binding of the SLP-76 SH2 domain to ADAP is critical for oligomerization of SLP-76 signaling complexes in stimulated T cells. Mol Cell Biol 33:4140-51
Ghirlando, Rodolfo; Balbo, Andrea; Piszczek, Grzegorz et al. (2013) Improving the thermal, radial, and temporal accuracy of the analytical ultracentrifuge through external references. Anal Biochem 440:81-95
Sousa, Alioscka A; Morgan, Jeffrey T; Brown, Patrick H et al. (2012) Synthesis, characterization, and direct intracellular imaging of ultrasmall and uniform glutathione-coated gold nanoparticles. Small 8:2277-86
Zhao, Huaying; Berger, Anthony J; Brown, Patrick H et al. (2012) Analysis of high-affinity assembly for AMPA receptor amino-terminal domains. J Gen Physiol 139:371-88
Zhao, Huaying; Brown, Patrick H; Schuck, Peter (2011) On the distribution of protein refractive index increments. Biophys J 100:2309-17

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