The family of glycogen phosphorylases: structure and function

Glycogen phosphorylase plays a central role in the mobilization of carbohydrate reserves in a wide variety of organisms and tissues. While rabbit muscle phosphorylase remains the most studied and best characterized of phosphorylases, recombinant DNA techniques have led to the recent appearance of primary sequence data for a wide variety of phosphorylase enzymes. The functional properties of rabbit muscle phosphorylases are reviewed and then compared to properties of phosphorylases from other tissues and organisms. Tissue expression patterns and the chromosomal localization of mammalian phosphorylases are described. Differences in functional properties among phosphorylases are related to new structural information. Evolutionary relationships among phosphorylases as afforded by comparative analysis of proteins and gene sequences are discussed.     

Photooxidation of alpha-glucan phosphorylases from rabbit muscle and potato tubers.

Photooxidation of alpha-glucan phosphorylases from rabbit muscle and potato tubers in the presence of rose bengal leads to a rapid loss of enzymatic activity which follows first-order kinetics. The process is pH dependent, being more rapid at higher pH. The inactivation is closely related to the destruction of histidine residues in the enzyme. It is suggested that histidine residues are largely responsible for the loss of enzymatic activity in the photooxidation. The inactivation of potato phosphorylase is retarded by substrates, whereas that of the muscle enzyme is not. The rate of photoinactivation of muscle phosphorylase b is increased with AMP, and decreased with ATP, ADP, IMP and glucose-6-P. This finding is considered to be closely related to the allosteric transition of phosphorylase.     

Complementation of subunits from glycogen phosphorylases of frog and rabbit skeletal muscle and rabbit liver.

Activity can be induced in potentially active rabbit skeletal muscle phosphorylase monomers covalently bound to Sepharose by noncovalent interaction with soluble subunits carrying inactive pyridoxal 5'-phosphate analogs or even salicyladlehyde. These analogs are themselves incapable of reconstituting active holophorphorylase from apophosphorylase. Phosphorylases with one intrinsically inactive and one potentially active subunit have about one half of the activity of the native phosphorylase dimer. The usefulness of this technique for subunit complementation was demonstrated by forming hybrid phosphorylases with inactive Sepharose-bound rabbit skeletal muscle subunits containing pyridoxal 5'-phosphate monomethylester and soluble activatable frog muscle and rabbit liver phosphorylase monomers. The inactive Sepharose-bound subunit induced in each case activity in the soluble subunit. But whereas the inactive rabbit muscle phosphorylase subunit even transmitted its characteristic temperature dependence of the rate of the reaction to the frog muscle subunit, it could not propagate its control properties to the liver enzyme. Differences of hybrid phosphorylases are related to immunological and amino acid divergencies among the component enzymes.     

Hemopexin: structure,function, and regulation

Hemopexin (HPX) is the plasma protein with the highest binding affinity to heme among known proteins. It is mainly expressed in liver, and belongs to acute phase reactants, the synthesis of which is induced after inflammation. Heme is potentially highly toxic because of its ability to intercalate into lipid membrane and to produce hydroxyl radicals. The binding strength between heme and HPX, and the presence of a specific heme-HPX receptor able to catabolize the complex and to induce intracellular antioxidant activities, suggest that hemopexin is the major vehicle for the transportation of heme in the plasma, thus preventing heme-mediated oxidative stress and heme-bound iron loss. In this review, we discuss the experimental data that support this view and show that the most important physiological role of HPX is to act as an antioxidant after blood heme overload, rather than to participate in iron metabolism. Particular attention is also put on the structure of the protein and on its regulation during the acute phase reaction.     

Affinity of glucose analogs for alpha-glucan phosphorylases from rabbit muscle and potato tubers

The action of phosphorylase b from rabbit muscle and potato phosphorylase was inhibited to various extents by several glucose analogs. Like glucose itself, all of the glucosidic oxygen-substituted analogs tested in kinetic experiments showed a nonlinear competitive inhibition for muscle phosphorylase b and a linear competitive one for potato phosphorylase. 5-Thio-D-glucose, one of the ring oxygen-substituted analogs, also inhibited the action of muscle phosphorylase b in the same manner, while the inhibition pattern of 5-amino-D-glucose (nojirimycin) was of a linear noncompetitive type. Since the conformation of 5-amino-D-glucose in aqueous solution is half-chair (Reese et al. (1971) Carbohyd. Res. 18, 381-388), the unusual kinetic behavior of the compound toward muscle phosphorylase b was supposed to be due to its half-chair conformation. In the glucosidic oxygen-substituted analogs, the affinity for both muscle phosphorylase b and potato phosphorylase decreased with decreasing order of magnitude of electronegativity of the glucosidic atom. The strong positive correlation between the affinity and the electronegativity suggests that D-glucose-1-P, the substrate, may bind to phosphorylase with the formation of a hydrogen bond between its glucosidic oxygen and a hydrogen donor of the enzyme.     

Comparative studies on muscle AMP-deaminase--I. Purification, molecular weight, subunit structure and metal content of the enzymes from rat, rabbit, hen, frog and pikeperch.

1. AMP-deaminase (EC 3.5.4.6) from skeletal muscle of frog and pikeperch was purified to homogeneity and compared with the homogeneous enzymes purified from rat, rabbit and hen skeletal muscle. 2. Their molecular weight was close to 280,000, every enzyme consisted of four identical subunits of molecular weight about 70,000. 3. All enzymes were found to contain about two atoms of zinc per molecule. 4. Minor differences of u.v.-absorption spectra between amphibian and fish muscle enzyme as compared with mammalian and bird muscle enzyme were found.     

Glucose transporters: structure, function, and regulation

Glucose is transported into the cell by facilitated diffusion via a family of structurally related proteins, whose expression is tissue-specific. One of these transporters, GLUT4, is expressed specifically in insulin-sensitive tissues. A possible change in the synthesis and/or in the amount of GLUT4 has therefore been studied in situations associated with an increase or a decrease in the effect of insulin on glucose transport. Chronic hyperinsulinemia in rats produces a hyper-response of white adipose tissue to insulin and resistance in skeletal muscle. The hyper-response of white adipose tissue is associated with an increase in GLUT4 mRNA and protein. In contrast, in skeletal muscle, a decrease in GLUT4 mRNA and a decrease (tibialis) or no change (diaphragm) in GLUT4 protein are measured, suggesting a divergent regulation by insulin of glucose transport and transporters in the 2 tissues. In rodents, brown adipose tissue is very sensitive to insulin. The response of this tissue to insulin is decreased in obese insulin-resistant fa/fa rats. Treatment with a beta-adrenergic agonist increases insulin-stimulated glucose transport, GLUT4 protein and mRNA. The data suggest that transporter synthesis can be modulated in vivo by insulin (muscle, white adipose tissue) or by catecholamines (brown adipose tissue).     

Biology of amyloid: structure, function, and regulation

Amyloids are highly ordered cross-β sheet protein aggregates associated with many diseases including Alzheimer's disease, but also with biological functions such as hormone storage. The cross-β sheet entity comprising an indefinitely repeating intermolecular β sheet motif is unique among protein folds. It grows by recruitment of the corresponding amyloid protein, while its repetitiveness can translate what would be a nonspecific activity as monomer into a potent one through cooperativity. Furthermore, the one-dimensional crystal-like repeat in the amyloid provides a structural framework for polymorphisms. This review summarizes the recent high-resolution structural studies of amyloid fibrils in light of their biological activities. We discuss how the unique properties of amyloids gives rise to many activities and further speculate about currently undocumented biological roles for the amyloid entity. In particular, we propose that amyloids could have existed in a prebiotic world, and may have been the first functional protein fold in living cells.     

Malic enzymes of higher plants: characteristics, regulation, and physiological function

The characteristics and distribution of the malic enzyme in plants is discussed as well as those features which appear to be limited to the plant NAD malic enzyme. Regulation of the malic enzyme as it relates to the physiological roles of this enzyme is also discussed.     

Dolichol-phosphate mannose synthase: structure, function and regulation

Glycosylation is the major modification of proteins, and alters their structures, functions and localizations. Glycosylation of secretory and surface proteins takes place in the endoplasmic reticulum and Golgi apparatus in eukaryotic cells and is classified into four modification pathways, namely N- and O-linked glycosylations, glycosylphosphatidylinositol (GPI)-anchor and C-mannosylation. These modifications are accomplished by sequential addition of single monosaccharides (O-linked glycosylation and C-mannosylation) or en bloc transfer of lipid-linked oligosaccharides (N-linked glycosylation and GPI) onto the proteins. The glycosyltransferases involved in these glycosylations are categorized into two classes based on the type of sugar donor, namely nucleotide-sugars and dolichol-phosphate-sugars, in which the sugar moiety is mannose or glucose. The sugar transfer from dolichol-phosphate-sugars occurs exclusively on the luminal side of the endoplasmic reticulum and is utilized in all four glycosylation pathways. In this review, we focus on the biosynthesis of dolichol-phosphate-mannose, and particularly on the mammalian enzyme complex involved in the reaction.     

The rabbit muscle phosphofructokinase gene. Implications for protein structure, function, and tissue specificity

Sequence homologies between bacterial and rabbit muscle phosphofructokinases and between the amino- and carboxyl-terminal halves of the latter suggest that the mammalian enzyme evolved from a prokaryotic progenitor by gene duplication and divergence (Poorman, R. A., Randolph, A., Kemp, R. G., and Heinrikson, R. L. (1984) Nature 309, 467-469). We have isolated the gene for the rabbit enzyme and determined the nucleotide sequence for all the exons and most of the introns. This represents the first eukaryotic phosphofructokinase gene ever sequenced. The cloned gene is 17 kilobase pairs long. The coding sequence for 780 amino acids is split into 22 exons ranging in size from 15 to 63 codons. Sequence analysis shows that 75% of the bases at the third position of the codons in these exons are either G or C. Exons XV and XVI code for the 30 amino acid residues which were left unidentified in the published primary structure for this enzyme. When overlaid on the structure of the protein, most of the introns are located between or near the ends of the secondary structural elements but not at analogous positions in the two protein-coding halves of the gene.