The crystallographic structure of the G protein heterotrimer Gi alpha 1(GDP)beta 1 gamma 2 (at 2.3 A) reveals two nonoverlapping regions of contact between alpha and beta, an extended interface between beta and nearly all of gamma, and limited interaction of alpha with gamma. The major alpha/beta interface covers switch II of alpha, and GTP-induced rearrangement of switch II causes subunit dissociation during signaling. Alterations in GDP binding in the heterotrimer (compared with alpha-GDP) explain stabilization of the inactive conformation of alpha by beta gamma. Repeated WD motifs in beta form a circularized sevenfold beta propeller. The conserved cores of these motifs are a scaffold for display of their more variable linkers on the exterior face of each propeller blade.

A fundamental goal in cellular signaling is to understand allosteric communication, the process by which signals originating at one site in a protein propagate reliably to affect distant functional sites. The general principles of protein structure that underlie this process remain unknown. Here, we describe a sequence-based statistical method for quantitatively mapping the global network of amino acid interactions in a protein. Application of this method for three structurally and functionally distinct protein families (G protein-coupled receptors, the chymotrypsin class of serine proteases and hemoglobins) reveals a surprisingly simple architecture for amino acid interactions in each protein family: a small subset of residues forms physically connected networks that link distant functional sites in the tertiary structure. Although small in number, residues comprising the network show excellent correlation with the large body of mechanistic data available for each family. The data suggest that evolutionarily conserved sparse networks of amino acid interactions represent structural motifs for allosteric communication in proteins.

Regulators of G protein signaling (RGS) proteins act as GTPase-activating proteins (GAPs) toward the ␣ subunits of heterotrimeric, signal-transducing G proteins. RGS11 contains a G protein ␥ subunit-like (GGL) domain between its Dishevelled͞Egl-10͞Pleckstrin and RGS domains. GGL domains are also found in RGS6, RGS7, RGS9, and the Caenorhabditis elegans protein EGL-10. Coexpression of RGS11 with different G ␤ subunits reveals specific interaction between RGS11 and G ␤5 . The expression of mRNA for RGS11 and G ␤5 in human tissues overlaps. The G ␤5 ͞RGS11 heterodimer acts as a GAP on G ␣o , apparently selectively. RGS proteins that contain GGL domains appear to act as GAPs for G ␣ proteins and form complexes with specific G ␤ subunits, adding to the combinatorial complexity of G protein-mediated signaling pathways.Proteins belonging to the RGS (regulators of G protein signaling) family constitute a newly appreciated group of at least 20 mammalian gene products that act as GTPaseactivating proteins (GAPs) on the ␣ subunits of heterotrimeric, signal-transducing G proteins (1-3). As such, RGS proteins can serve as negative regulators of G proteinmediated signaling pathways by speeding the inactivation of GTP-bound G ␣ subunits. Although several members of the RGS family are relatively simple Ϸ25 kDa proteins that contain little more than a characteristic RGS domain, others include modules that impart additional functions. For example, RGS12 can associate in vitro with certain G protein-coupled receptors by virtue of an alternatively spliced PDZ (PSD-95͞ Dlg͞Z0-1) domain (4), and p115, a guanine nucleotide exchange factor for the low-molecular-weight GTPase rho, contains an RGS domain that imparts sensitivity to regulation by G protein ␣ subunits (5, 6).We describe here a novel G protein ␥ subunit-like domain (GGL; pronounced giggle) that is found in several mammalian RGS proteins (RGS6, RGS7, RGS9, and RGS11) and in EGL-10, an RGS protein of Caenorhabditis elegans. The GGL domains of RGS11 and RGS7 interact preferentially with the G protein ␤ 5 subunit, and the complex of RGS11 and ␤ 5 has GAP activity toward the G protein ␣ o subunit. MATERIALS AND METHODSGeneration of Expression Constructs. cDNAs for RGS11 and various G protein subunits were cloned from human brain or retinal mRNA, from mouse retinal mRNA, or were obtained as described (7,8); all amplified cDNAs were verified by sequencing. Human RGS7 cDNA was a kind gift of Paul F. Worley (Johns Hopkins University). cDNAs encoding G protein subunits were subcloned into the mammalian expression vector pcDNA3.1-Zeo (Invitrogen), and G ␥ and RGS protein cDNAs were subcloned in-frame with an N-terminal tandem hemagglutinin (HA)-epitope tag into a modified pcDNA3.1 vector. Recombinant baculoviruses expressing native or hexahistidine-tagged RGS11 or G ␤5 subunits were generated by using the Bac-To-Bac system by following the manufacturer's protocols (Life Technologies, Gaithersburg, MD).In Vitro Transcription and Translation. Reactions were performed using the T...

The branching ratio of core-valence transitions in electron energy-loss spectroscopy and x-ray absorption spectroscopy is linearly related to the expectation value of the spin-orbit operator of the valence states. Here, we analyze the branching ratio of the N(4,5) edges in the actinides and find that the spin-orbit sum rule gives an accurate result without the need to include the core-valence interactions. The branching ratio is not only useful to study the variations in the 5f spin-orbit interaction, it also allows us to constrain the 5f count for given angular-momentum coupling conditions.