Cooperativity and noncooperativity in the binding of NAD analogs to rabbit muscle glyceraldehyde-3-phosphate dehydrogenase

D, Eby; Me, Kirtly

doi:10.1021/bi00655a021

Cited by 21 publications

(10 citation statements)

References 17 publications

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“…Such a phenomenon can be encountered only if the competing ligand binds noncooperatively and, in addition, if its binding is not affecting (and not affected by) the binding of ENAD+ to neighboring subunits (21); These results confirm previous findings that ADP-Rib binds noncooperatively to Gra-P dehydrogenase (36).…”

Section: Resultssupporting

confidence: 88%

Mechanism of negative cooperativity in glyceraldehyde-3-phosphate dehydrogenase deduced from ligand competition experiments.

Henis¹,

Levitzki²

1980

Proc. Natl. Acad. Sci. U.S.A.

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It is shown that the modulation in the negative cooperativity of ligand binding by another, competing ligand that binds noncooperatively is accounted for exclusively by the ligand-induced sequential model. It is therefore suggested that whenever such a phenomenon is observed it argues strongly in favor of the sequential model. The advantages and limitations of this approach are evaluated. The binding of the coenzymes NAD+ and nicotinamide-1-N6-ethenoadenine dinucleotide to rabbit muscle apo-glyceraldehydephosphate dehydrogenase [D-glyceraldehyde-3-phosphate:NAD+ oxidoreductase (phosphorylating; EC 1.2.1.12] exhibits strong negative cooperativity, whereas acetylpyridine adenine dinucleotide, ATP, and ADPribose bind noncooperatively to the NAD+ sites. The strong negative cooperativity in coenzyme binding was found to be abolished in the presence of acetylpyridine adenine dinucleotide and strongly weakened by ATP, ADP, and AMP, but was not affected by addition of ADP-ribose. These findings demonstrate that the negative cooperativity in coenzyme binding to this enzyme results from sequential conformational changes and exclude the pre-existent asymmetry model as a possible explanation. These results also support the view that the structure of the pyridine moiety of the coenzyme analogs plays a role in orienting the adenine moiety at the adenine subsite, therefore affecting the cooperativity in the binding of the coenzyme analog which is mediated through the adenine subsites.

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Section: Resultssupporting

confidence: 88%

Mechanism of negative cooperativity in glyceraldehyde-3-phosphate dehydrogenase deduced from ligand competition experiments.

Henis¹,

Levitzki²

1980

Proc. Natl. Acad. Sci. U.S.A.

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“…Catalytic activity of the D302A‐R406A complementation mutant, which has only one functional active site, is comparable to that of the wild‐type enzyme (Figure 6), supporting the structural observation that only one site of the homodimer is catalytically competent at a time. Mechanisms of half‐of‐sites reactivity have been previously observed in enzymes such as adenylyl transferases (Izard and Geerlof, 1999), tyrosyl‐tRNA synthetase (First and Fersht, 1993) and dehydrogenases (Eby and Kirtly, 1976; Nichols et al , 2004), as well as ligand‐binding proteins like the aspartate receptors (Biemann and Koshland, 1994). However, unlike these proteins, where half‐of‐sites‐reactivity results in negative cooperativity, Rv1900c shows slight positive cooperativity for product formation with respect to ATP.…”

Section: Discussionmentioning

confidence: 99%

Origin of asymmetry in adenylyl cyclases: structures of Mycobacterium tuberculosis Rv1900c

Sinha

Wetterer

Sprang

et al. 2005

EMBO J

108

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Rv1900c, a Mycobacterium tuberculosis adenylyl cyclase, is composed of an N-terminal alpha/beta-hydrolase domain and a C-terminal cyclase homology domain. It has an unusual 7% guanylyl cyclase side-activity. A canonical substrate-defining lysine and a catalytic asparagine indispensable for mammalian adenylyl cyclase activity correspond to N342 and H402 in Rv1900c. Mutagenic analysis indicates that these residues are dispensable for activity of Rv1900c. Structures of the cyclase homology domain, solved to 2.4 A both with and without an ATP analog, form isologous, but asymmetric homodimers. The noncanonical N342 and H402 do not interact with the substrate. Subunits of the unliganded open dimer move substantially upon binding substrate, forming a closed dimer similar to the mammalian cyclase heterodimers, in which one interfacial active site is occupied and the quasi-dyad-related active site is occluded. This asymmetry indicates that both active sites cannot simultaneously be catalytically active. Such a mechanism of half-of-sites-reactivity suggests that mammalian heterodimeric adenylyl cyclases may have evolved from gene duplication of a primitive prokaryote-type cyclase, followed by loss of function in one active site.

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“…The fluorescent coenzyme analogue eNAD41 has been used by several workers to probe the adenine binding sites of NAD4-linked dehydrogenases as well as the conformation of the bound coenzyme analogue (Lee & Everse, 1973;Schlessinger & Levitzki, 1974;Schlessinger et al, 1975;Luisi et al, 1975;Gafni, 1977). The pronounced enhancement of fluorescence of eNAD4 upon binding was used to follow the binding mechanism and to determine the dissociation constants.…”

mentioning

confidence: 99%

“…The pronounced enhancement of fluorescence of eNAD4 upon binding was used to follow the binding mechanism and to determine the dissociation constants. Free eNAD4, in aqueous solution, was found to be in an equilibrium between folded and open conformations (Lee & Everse, 1973; Gruber & Leonard, 1975;Gafni, 1977). In the folded conformation partial quenching of the fluorescence is caused by stacking of the ethenoadenine and nicotinamide rings.…”

mentioning

confidence: 99%

Accessibility of adenine binding sites in dehydrogenases to small molecules studied by fluorescence quenching

Gafni¹

1979

Biochemistry

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Quenching of the fluorescence of ethenoadenine derivatives by iodide ions and by methionine was studied in solution and when the nucleotides were bound to several dehydrogenases. The fluorescence of epsilonADPR in neutral aqueous solution is dynamically quenched by both quenching agents. The quenching of free epsilonNAD+ by methionine was found to be predominantly static and was satisfactorily described to result from complex formation between quencher and dinucleotide. The rat constant for quenching by iodide of epsilonNAD+ in the ternary complex with LADH and pyrazole is comparable to that of free epsilonADPR or epsilonADP. it is concluded that the bound epsilon-adenine ring is partially exposed to the solvent. The opening, to the solvent, of the adenine binding site is not large enough to allow free methionine diffusion since the rate constant for quenching of bound coenzyme by this quenching agent is relatively small. The difference between the rate constants for quenching of free and enzyme bound nucleotide was used to evaluate the binding constants of epsilonADPR to GPDH, epsilonNAD+ to LDH, and oxalate to the LDH:epsilonNAD+ complex. This technique may prove to be particularly useful when the binding of a fluorescent ligand to a protein is not accompanied by significant changes in its fluorescence.

show abstract

Cooperativity and noncooperativity in the binding of NAD analogs to rabbit muscle glyceraldehyde-3-phosphate dehydrogenase

Cited by 21 publications

References 17 publications

Mechanism of negative cooperativity in glyceraldehyde-3-phosphate dehydrogenase deduced from ligand competition experiments.

Mechanism of negative cooperativity in glyceraldehyde-3-phosphate dehydrogenase deduced from ligand competition experiments.

Origin of asymmetry in adenylyl cyclases: structures of Mycobacterium tuberculosis Rv1900c

Accessibility of adenine binding sites in dehydrogenases to small molecules studied by fluorescence quenching

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