In this issue

1775 Quantitative in vivo solubility and reconstitution of truncated circular permutants of green fluorescent protein

Yao-Ming Huang, Sasmita Nayak, Christopher Bystroff

Green fluorescent protein can be split into two parts that sometimes reassemble to reconstitute fluorescence. Here, Huang, Nayak, and Bystroff exhaustively removed each of the 11 beta strands and the central helix from the GFP sequence and then measured the solubility in bacteria. By co-expressing each LOO-GFP with its left-out segment, the authors measured the ability of the LOO-GFPs to bind their missing piece and glow. What was found is that the solubility of the LOO-GFP depends very much on which part of the molecule was left out. Some parts of the protein are more essential to efficient folding than others.

Illustration 1.

1790 A tale of two GTPases in cotranslational protein targeting

Ishu Saraogi, David Akopian, Shu-Ou Shan

The ‘GTPase switch’ paradigm, in which a GTPase switches between a GTP-bound ‘on’ state and a GDP-bound ‘off’ state, has dominated our understanding of small GTPases for over two decades. Nevertheless, this classical paradigm could not explain the action of a growing class of GTPases that undergo nucleotide-dependent dimerization cycles. The work of Saraogi, Akopian, and Shan in the SRP and SRP receptor reveals a distinct mode of regulation by this new class of GTPases. The results suggest that, instead of the classic ‘bi-modal’ switch, these GTPases can undergo multiple and discrete conformational changes during their dimerization and activation. These ‘multi-state’ regulatory GTPases may be particularly suitable to drive cyclic processes where multiple factors must bind and later dissociate in a sequential and highly coordinated manner, allowing these complex cellular pathways to achieve both high efficiency and exquisite specificity.

Illustration 2.

1781 Surviving the sun: Repair and bypass of DNA UV lesions

Wei Yang

Each cell devotes a large number of proteins to maintain genome stability and repair a wide variety of damaged DNA bases. The first step in repair is to locate lesions and match damaged bases with appropriate repair proteins for restoration. A common feature of damaged bases, including ultraviolet-induced DNA lesions, is reduced local stability of the double helix. At lesion sites, DNA easily bends, unwinds and separates, and returns to a canonical double helix after distortion is impaired. Here, Yang hypothesizes that DNA-repair proteins use the energy of ATP hydrolysis to facilitate lesion recognition by checking hysteresis of DNA distortion.

Illustration 3.

1876 An approach to crystallizing proteins by metal-mediated synthetic symmetrization

Arthur Laganowsky, Minglei Zhao, Angela B. Soriaga, Michael R. Sawaya, Duilio Cascio, Todd O. Yeates

New methods continue to develop for engineering proteins to improve their chances of being crystallized. In one line of attack, a protein of interest is engineered to form a series of distinct oligomeric arrangements, each of which presents unique opportunities for forming a crystal lattice. In the present work, partial metal binding sites (e.g. two histidine residues on the side of an alpha helix) are engineered into different spots on the surface of a protein. Addition of metals then leads to a variety of different oligomeric forms of the protein, including dimers, trimers, and tetramers. The method showed promise in experiments on model protein systems, where numerous new crystal forms were obtained.

Illustration 4.