Allosteric mechanisms can be distinguished using structural mass spectrometry.

The activity of many proteins, including metabolic enzymes, molecular machines, and ion channels, is often regulated by conformational changes that are induced or stabilized by ligand binding. In cases of multimeric proteins, such allosteric regulation has often been described by the concerted Monod-Wyman-Changeux and sequential Koshland-Némethy-Filmer classic models of cooperativity. Despite the important functional implications of the mechanism of cooperativity, it has been impossible in many cases to distinguish between these various allosteric models using ensemble measurements of ligand binding in bulk protein solutions.

The dimeric structure of the Cpn60.2 chaperonin of Mycobacterium tuberculosis at 2.8 Å reveals possible modes of function.

Mycobacterium tuberculosis expresses two proteins (Cpn60.1 and Cpn60.2) that belong to the chaperonin (Cpn) family of heat shock proteins.

A method for removing effects of nonspecific binding on the distribution of binding stoichiometries: application to mass spectroscopy data.

There is often an interest in knowing, for a given ligand concentration, how many protein molecules have one, two, three, etc. ligands bound in a specific manner. This is a question that cannot be addressed using conventional ensemble techniques.

ATP-induced allostery in the eukaryotic chaperonin CCT is abolished by the mutation G345D in CCT4 that renders yeast temperature-sensitive for growth.

Saccharomyces cerevisiae yeast cells containing the chaperonin CCT (chaperonin-containing t-complex polypeptide 1 (TCP-1)) with the G345D mutation in subunit CCT4 (anc2-1) are temperature-sensitive for growth and display defects in organization of actin structure, budding and cell shape. In this first structure-function analysis of CCT, we show that this mutation abolishes both intra- and inter-ring cooperativity in ATP binding by CCT. The finding that a single mutation in only one subunit in each CCT ring has such drastic effects highlights the importance of allostery for its in vivo function.

Glu257 in GroEL is a sensor involved in coupling polypeptide substrate binding to stimulation of ATP hydrolysis.

The ATPase activity of many types of molecular chaperones is stimulated by polypeptide substrate binding via molecular mechanisms that are, for the most part, unknown. Here, we report that such stimulation of the ATPase activity of GroEL is abolished when its conserved apical domain residue Glu257 is replaced by alanine. This mutation is also found to convert the ATPase profile of GroEL, a group I chaperonin, into one that is characteristic of group II chaperonins.

Sequential ATP-induced allosteric transitions of the cytoplasmic chaperonin containing TCP-1 revealed by EM analysis.

The eukaryotic cytoplasmic chaperonin containing TCP-1 (CCT) is a hetero-oligomeric complex that assists the folding of actins, tubulins and other proteins in an ATP-dependent manner. To understand the allosteric transitions that occur during the functional cycle of CCT, we imaged the chaperonin complex in the presence of different ATP concentrations. Labeling by monoclonal antibodies that bind specifically to the CCTalpha and CCTdelta subunits enabled alignment of all the CCT subunits of a given type in different particles.