The Subunits

The cDNA encoding the /336 subunit of bovine T [19,20] and that of a human liver Gs/Gj [21] have been cloned. The two cDNAs encode for the identical polypeptide of 340 amino acids with a calculated Mr 37375. Polyclonal antibodies raised against 

As pointed out earlier in this article, T differs from other G proteins in that it is a peripheral membrane protein. After activation by Rho it seems to undergo subunit dissociation in which both its a subunit and its /3y complex dissociate from the Rho-containing membranes. Purification of brain G-proteins has shown that free a subunits of G0 and Gj are also water soluble, remaining in solution in the absence of detergents [74], The hydrophobicity of the whole ajSy G and Gj complexes was shown to be due to their j8y complexes 189]. Indeed, purified a subunits associate with phospholipid vesicles only if j8y complexes have been incorporated during vesicle formation [189]. Since the amino acid composition of T-/3 is equal to that of other G-j8s, but their ys differ, it follows that the principal role of y subunits should be to anchor non-T G proteins to the plasma membranes. This conclusion assumed, of course, that j8 subunits are not post-translationally modified in a tissue specific manner such that that they become water soluble in retinal photoreceptor cells and 

Effects of inhibitory receptors on K+ channels in tissues other than heart atria 

In GH cells SST inhibits prolactin secretion induced by cAMP analogs and K+-induced depolarization in addition to inhibiting adenylyl cyclase activity [192], Similar data were also obtained with ACTH-secreting AtT-20 cells [203]. PTX blocks SST inhibition of ligand-induced and cAMP analog-induced secretions in both GH and AtT-20 cells [106,107]. Thus, it is clear that a G protein is involved in both the cAMP dependent and the cAMP-independent actions of SST. Quin-2 measurements showed that exposure of cells to SST lowers the intracellular Ca2+ levels. This effect is also PTX-sensitive [211-213], The decline in intracellular Ca2+ was concluded to be secondary to SST hyperpolarizing the cells by increasing K+ conductance [214,215]. Acetylcholine, like SST, causes inhibition of adenylyl cyclase, hy- 

Some four years ago studies on erythrocuprein which was oxidized with mild performic acid or reduced and alkylated revealed that the protein contains a subunit of molecular weight about 12,000 (129). Subunits of molecular weight 16,000 were detected after the native erythrocuprein had been subjected to sodium dodecyl sulphate treatment in the presence of mercaptoethanol (74, 82). In addition to the 16,000 molecular-weight subunits, it was noted that a short-time treatment revealed protein portions of molecular weight of approximately 64,000 which may be attributed to tetrameric species (Fig. 23). 

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The Fe-N mode is at 222 in the R state and 207 cnY in the T state for the a subunits, but only shifted to 218 T state for the (3 subunits. This is consistent with the interpretation that the Fe-imidazole interations are weakened more in the T state of the a subunits than p subunits. Time-resolved resonance Raman studies have shown that the R T switch is complete on a 10 ps tuuescale [38]. Finally, UV excitation of the aromatic protein side chains yields... 

Indole, the product of the a-reaction and the reactant for the /3-reaction, is passed directly from the a-subunit to the /3-subunit and cannot be detected as a free intermediate. 

Meighen, E. A., and Bartlet, I. (1980). Complementation of subunits from different bacterial luciferases. Evidence for the role of the (3 subunit in the bioluminescent mechanism. J. Biol. Chem. 255 11181— 11187. 

The a subunits, for which two isoforms exist in mammals (al, a2), contain conventional protein serine/threonine kinase domains at the N-terminus, with a threonine residue in the activation loop (Thr-172) that must be phosphorylated by upstream kinases (see below) before the kinase is active. The kinase domain is followed by an autoinhibitory domain, whose effect is somehow relieved by interaction with the other subunits. The C-terminal domain of the a subunit is required for the formation of a complex with the C-terminal domain of the (3 subunit, which in turn mediates binding to the y subunit. The al and a2 catalytic subunit isoforms are widely distributed, although a2 is most abundant in muscle and may be absent in cells of the endothelial/hemopoietic lineage. 

Other examples include rifampin resistance due to mutations in the ipoB gene encoding the (3-subunit of RNA polymerase, or oxazolidinone resistance due to a G2576T mutation in the gene for the 23 S rRNA as central part of the 50S large ribosomal subunit. Macrolide resistance is based upon the alteration of nucleotide A2058 by a point mutation. 

Insulin Receptor. Figure 1 Structure and function of the insulin receptor. Binding of insulin to the a-subunits (yellow) leads to activation of the intracellular tyrosine kinase ((3-subunit) by autophosphorylation. The insulin receptor substrates (IRS) bind via a phospho-tyrosine binding domain to phosphorylated tyrosine residues in the juxtamembrane domain of the (3-subunit. The receptor tyrosine kinase then phosphorylates specific tyrosine motifs (YMxM) within the IRS. These tyrosine phosphorylated motifs serve as docking sites for some adaptor proteins with SRC homology 2 (SH2) domains like the regulatory subunit of PI 3-kinase. 

One amino acid substitution within a specific region in the (3-subunit of the RNA-polymerase, the target molecule of lifampicin, is necessary to establish high-level resistance to this antibiotic. 

Histidine phosphatases and aspartate phosphatases are well established in lower organisms, mainly in bacteria and in context with two-component-systems . Reversible phosphorylation of histidine residues in vertebrates is in its infancy. The first protein histidine phosphatase (PHP) from mammalian origin was identified just recently. The soluble 14 kD protein does not resemble any of the other phosphatases. ATP-citrate lyase and the (3-subunit of heterotrimeric GTP-binding proteins are substrates of PHP thus touching both, metabolic pathways and signal transduction [4]. 

The 3 subunits ((31 -(34) are membrane proteins with a single transmembrane domain and an extracellular immunoglobulin-like motif, and perform the regulatory roles of the sodium channel. The (31 subunit accelerates the activation and inactivation kinetics. The (32 subunit is covalently linked to the a subunit, and is necessary for the efficient assembly of the channel. The more recently identified (33 subunit is homologous to (31, but differs in its distribution within the brain and in a weaker accelerating property. The (34 subunit is similar to (32 and is covalently linked to the a subunit. 

This key enzyme of the dissimilatory sulfate reduction was isolated from all Desulfovibrio strains studied until now 135), and from some sulfur oxidizing bacteria and thermophilic Archaea 136, 137). The enzymes isolated from sulfate-reducing bacteria contain two [4Fe-4S] clusters and a flavin group (FAD) as demonstrated by visible, EPR, and Mossbauer spectroscopies. With a total molecular mass ranging from 150 to 220 kDa, APS reductases have a subunit composition of the type 012)32 or 02)3. The subunit molecular mass is approximately 70 and 20 kDa for the a and )3 subunits, respectively. Amino-acid sequence data suggest that both iron-sulfur clusters are located in the (3 subunit... 

Bisphosphoglycerate (BPG) in the central cavity of deoxyHb forms salt bonds with the (3 subunits that stabilize deoxyHb. On oxygenation, the central cavity contracts, BPG is extruded, and the quaternary structure loosens. 

Figure 12-9. Mechanism of ATP production by ATP synthase. The enzyme compiex consists of an Fq sub-compiex which is a diskof "C" protein subunits. Attached is a y-subunit in the form of a "bentaxie." Protons passing through the disk of "C" units cause it and the attached y-subunit to rotate. The y-subunit fits inside the F, subcompiex of three a- and three (3-sub-units, which are fixed to the membrane and do not rotate. ADP and P are taken up sequentiaiiy by the (3-subunits to form ATP, which is expeiied as the rotating y-subunit squeezes each (3-subunit in turn. Thus, three ATP moiecuies are generated per revoiution. For ciarity, not aii the subunits that have been identified are shown—eg, the "axie" aiso contains an e-subunit.

The process of RNA synthesis in bacteria—depicted in Figure 37-3—involves first the binding of the RNA holopolymerase molecule to the template at the promoter site to form a PIC. Binding is followed by a conformational change of the RNAP, and the first nucleotide (almost always a purine) then associates with the initiation site on the 3 subunit of the enzyme. In the presence of the appropriate nucleotide, the RNAP catalyzes the formation of a phosphodiester bond, and the nascent chain is now attached to the polymerization site on the P subunit of RNAP. (The analogy to the A and P sites on the ribosome should be noted see Figure... 

The action of rifampicin is upon the /3 subunit of RNA polymerase. Binding of just one molecule of rifampicin inhibits the initiation stage of transeription in whieh the first nucleotide is incorporated in the RNA ehain. Once started, transeription itself is not inhibited. It has been suggested that the stmeture of rifampiein resembles that of two adenosine nucleotides in RNA this may form the basis of the binding of the antibiotic to the j3 subunit. One problem is the rapid development of resistanee in organisms due... 

The NHase responsible for aldoxime metabolism from the i -pyridine-3-aldoxime-degrading bacterium, Rhodococcus sp. strain YH3-3, was purified and characterized. Addition of cobalt ion was necessary for the formation of enzyme. The native enzyme had a Mr of 130000 and consisted of two subunits (a-subunit, 27 100 (3-subunit, 34500). The enzyme contained approximately 2 mol cobalt per mol enzyme. The enzyme had a wide substrate specificity it acted on aliphatic saturated and unsaturated as well as aromatic nitriles. The N-terminus of the (3-subunit showed good sequence similarities with those of other NHases. Thus, this NHase is part of the metabolic pathway for aldoximes in microorganisms. 

Pihlajaniemi, T., Helaakoski, T., Tasanen, K., Myllyla, R., Huhtal, M.-L., Koivu, J.K. and Kivirikko, K.I. (1987) Molecular cloning of the 3-subunit of human prolyl 4-hydroxylase. This subunit and protein disulphide isomerase are products of the same gene. EMBOJournals, 643-649. 

In the first family, the metal is coordinated by one molecule of the pterin cofactor, while in the second, it is coordinated to two pterin molecules (both in the guanine dinucleotide form, with the two dinucleotides extending from the active site in opposite directions). Some enzymes also contain FejSj clusters (one or more), which do not seem to be directly linked to the Mo centers. The molybdenum hydroxylases invariably possess redox-active sites in addition to the molybdenum center and are found with two basic types of polypeptide architecture. The enzymes metabolizing quinoline-related compounds, and derivatives of nicotinic acid form a separate groups, in which each of the redox active centers are found in separate subunits. Those enzymes possessing flavin subunits are organized as a2jS2A2, with a pair of 2Fe-2S centers in the (3 subunit, the flavin in the (3 subunit, and the molybdenum in the y subunit. 

To establish whether rifaximin, like the other members of the rifamycin family [36, 58], specifically inhibits bacterial RNA synthesis the effect of this antibiotic as well as that of rifampicin and chloramphenicol on RNA (via 3H-uridine incorporation), DNA (via 3H-thymidine incorporation) and protein (via 35S-methionine incorporation) synthesis was studied in growing cultures of Escherichia coli [59], While chloramphenicol reduced protein synthesis, both rifaximin and rifampicin inhibited RNA synthesis in a concentration-dependent fashion. In contrast, none of them affected 3H-thymidine incorporation into DNA. These data suggest that rifaximin, like rifampicin, inhibits RNA synthesis by binding the (3 subunit of the bacterial DNA-dependent RNA polymerase [60],... 

Figure 11.2 Structure of the insulin receptor (a). Binding of insulin promotes autophosphorylation of the (3-subunits, where each (3-subunit phosphorylates the other (3-subunit. Phosphate groups are attached to three specific tyrosine residues (tyrosines 1158, 1162 and 1163), as indicated in (b). Activation of the (3-subunit s tyrosine kinase activity in turn results in the phosphorylation of various intracellular (protein) substrates which trigger the mitogen-activated protein kinase and/or the phosphoinositide (PI-3) kinase pathway responsible for inducing insulin s mitogenic and metabolic effects. The underlying molecular events occurring in these pathways are complex (e.g. refer to Combettes-Souverain, M. and Issad, T. 1998. Molecular basis of insulin action. Diabetes and Metabolism, 24, 477-489)...

Fig. 3. Three-dimensional structures of three examples of superstructures formed by sequence repeats a linear rod (the spectrin a-chain dimer [PDB 2spc]), a superhelix of repeats (armadillo repeats of importin a-subunit [PDB lbk5]), and a closed /3-propeller (WD40 repeats from a fragment of the /3-subunit of the guanine nucleotide binding protein 1 [PDB lgg2 chain B]).

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