• Molecules and Cells
• /
• v.26 no.1
• /
• pp.61-66
• /
• 2008

Cyclic AMP receptor protein (CRP) is allosterically activated by cAMP and functions as a global transcription regulator in enteric bacteria. Structural information on CRP in the absence of cAMP (apo-CRP) is essential to fully understand its allosteric behavior. In this study we demonstrated interdomain interactions in apo-CRP, using a comparative thermodynamic approach to the intact protein and its isolated domains, which were prepared either by limited proteolysis or using recombinant DNA. Thermal denaturation of the intact apo-CRP, monitored by differential scanning calorimetry, revealed an apparently single cooperative transition with a slight asymmetry. Combined with circular dichroism and fluorescence analysis, the thermal denaturation of apo-CRP could be interpreted as a coupled process involving two individual transitions, each attributable to a structural domain. When isolated individually, both of the domains exhibited significantly altered thermal behavior, thus pointing to the existence of non-covalent interdomain interactions in the intact apo-CRP. These observations suggest that the allosteric conformational change of CRP upon binding to cAMP is achieved by perturbing or modifying pre-existing interdomain interactions. They also underline the effectiveness of a comparative approach using calorimetric and structural probes for studying the thermodynamics of a protein.

• Journal of Photoscience
• /
• v.7 no.3
• /
• pp.79-86
• /
• 2000

Phytochromes are red and far-red light absorbing photoreceptors for photomorphogenesis in plants. The red/far wavelength reversible biliproteins are made up of two structural domains. The light-perceiving function of the photoreceptor resides in the N-terminal domain, whereas the signal transducing regulatory function is located within the C-terminal domain. The characteristic role of the phytochromes as phtosensory molecular switches is derived from the phototransformation between two distinct spectral forms, the red light absorbing Pr and the far-red light absorbing Pfr forms. The photoinduced Pr Pfr phototransformation accompanies subtle conformational changes throughout the phytochrome molecule. The conformational signals are subsequently transmitted to the C-terminal domain through various inter-domain crosstalks and induce the interaction of the activated C-terminal domain with phytochrome interacting factors. Thus the inter-domain crosstalks play critical roles in the photoactivation of the phytochromes. Posttranslational modifications, such as the phosphorylation of Ser-598, are also involved in this process through conformational changes and by modulating inter-domain signaling.

• Journal of Microbiology and Biotechnology
• /
• v.26 no.4
• /
• pp.637-647
• /
• 2016

Proliferating cell nuclear antigen (PCNA) is a critical eukaryotic replication accessory factor that supports DNA binding in DNA processing, such as DNA replication, repair, and recombination. PCNA consists of three toroidal-shaped monomers that encircle double-stranded DNA. The diverse functions of PCNA may be regulated by its interactions with partner proteins. Many of the PCNA partner proteins generally have a conserved PCNA-interacting peptide (PIP) motif, located at the N- or C- terminal region. The PIP motif forms a 3₁₀ helix that enters into the hydrophobic groove produced by an interdomain-connecting loop, a central loop, and a C-terminal tail in the PCNA. Post-translational modification of PCNA also plays a critical role in regulation of its function and binding partner proteins. Structural and biochemical studies of PCNA-protein will be useful in designing therapeutic agents, as well as estimating the outcome of anticancer drug development. This review summarizes the characterization of eukaryotic PCNA in relation to the protein structures, functions, and modifications, and interaction with proteins.