The folding and mutual interaction of the domains of yeast 3‐phosphoglycerate kinase

Adams, Benjamin; Burgess, Rowland J.; Pain, Roger H.

doi:10.1111/j.1432-1033.1985.tb09252.x

Cited by 42 publications

(31 citation statements)

References 23 publications

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“…In the yeast enzyme a large difference between the calculated and the observed concentration dependence of AG has been reported (a ratio 3: 1 for mcal/mobs) [37]. A ratio of these parameters greater than unity is considered evidence for a significant population of a folding intermediate [40] and it has been suggested that in PGK this species arises through the independent unfolding of one of its domains.…”

Section: Gdnhci Denaturationmentioning

confidence: 99%

“…A ratio of these parameters greater than unity is considered evidence for a significant population of a folding intermediate [40] and it has been suggested that in PGK this species arises through the independent unfolding of one of its domains. In the light of the fluorescence transitions observed for PGK [37,38] it would seem likely that the N domain would partially or completely unfold before the more stable C domain. Differentiating the polynomial described by Adams et al [37] using the same fractional exposure of residues as these authors to correct AG, we obtain the values of mca, listed in Table 2.…”

Section: Gdnhci Denaturationmentioning

confidence: 99%

“…In the light of the fluorescence transitions observed for PGK [37,38] it would seem likely that the N domain would partially or completely unfold before the more stable C domain. Differentiating the polynomial described by Adams et al [37] using the same fractional exposure of residues as these authors to correct AG, we obtain the values of mca, listed in Table 2. These calculated terms are all approximately three times greater than the observed values.…”

Section: Gdnhci Denaturationmentioning

confidence: 99%

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A comparison of the reactivity and stability of wild type and His388 Gln mutant phosphoglycerate kinase from yeast

Johnson

Cooper

Brown

1991

European Journal of Biochemistry

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A variety of physico-chemical techniques have been used to probe the possible interactions between the characteristic structural domains of yeast phosphoglycerate kinase by comparison of the wildtype enzyme with the specific H388Q mutant in which a potential interaction between His388 and Glu190 in the crucial interdomain region is disrupted. Enzyme kinetic studies indicate that, despite being structurally remote from the active site, this mutation has significant effects on both the VmaX and K,,, values for various substrates. The single cysteine residue in the N domain of the protein is markedly more reactive in the mutant, and this enhanced accessibility is moderated by binding of substrates and various anions. Differences are also observed in the near-ultraviolet CD spectra of these proteins. The chemical and thermal stability of the mutant enzyme is reduced, as indicated from guanidinium chloride and differential-scanning calorimetry denaturation studies. Moreover, interdomain interactions seem to be altered in the mutant, resulting in the appearance of independent thermal transition for the two domains, in contrast to the single cooperative transition observed for the wild-type enzyme. The conformational and/or dynamic effects of the mutation on the H388Q enzyme are therefore various and not solely localised in the hinge region.Phosphoglycerate kinase (PGK) catalyses reversible phosphoryl transfer from 1,3-bisphosphoglycerate to ADP in a key ATP-producing step of glycolysis. The enzyme reaction requires metal ions (Mg2+ or Mn'') which form a complex with the nucleotide substrates [I]. PGK isolated from various sources show strong sequence homology and are monomeric proteins with a molecular mass of about 45 kDa. The X-ray structures of the yeast and horse muscle enzymes [2, 31 have shown that the single polypeptide chain is organised into two structurally distinct domains of approximately equal size. In the yeast enzyme the nucleotide and glycerate substrates bind in separate sites close to the interdomain cleft. Mg-ATP or Mg-ADP molecules bind competitively in a shallow depression on the C domain while the most plausible site for 3-phosphoglycerate and 1,3-bisphosphoglycerate is located close to the face of the N domain [2]. In view of the distance between these binding sites (some 1 nm), a hinge-bending model has been proposed for PGK in which movement around the waist region (or hinge) between the two domains expels water and brings the substrates into an orientation and proximity which facilitate phosphoryl transfer [2, 31. Evidence for such a conformational change during catalysis has come from a number of studies including low angle X-ray scattering [4,5] [XI, while consistent with significant conformational change, has not been reproduced elsewhere [9]). Neither the extent nor the possible mechanism of hinge bending during catalysis are known, but the involvement of a His388-Glul90 interaction in the waist region of the enzyme as a mediator of domain closure has been proposed [ZJ. His388 is con...

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Section: Gdnhci Denaturationmentioning

confidence: 99%

Section: Gdnhci Denaturationmentioning

confidence: 99%

Section: Gdnhci Denaturationmentioning

confidence: 99%

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A comparison of the reactivity and stability of wild type and His388 Gln mutant phosphoglycerate kinase from yeast

Johnson

Cooper

Brown

1991

European Journal of Biochemistry

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“…The protein concentration was determined spectrophotometrically, using All ~ = 4.95 at 280nm (Adams, cm Burgess & Pain, 1985). subtraction of the normalized background (Guinier & Fournet, 1955):…”

Section: Methodsmentioning

confidence: 99%

Improved Signal-to-Background Ratio in Small-Angle X-ray Scattering Experiments with Synchrotron Radiation using an Evacuated Cell for Solutions

Dubuisson¹,

Décamps²,

Vachette³

1997

J Appl Cryst

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An evacuated, temperature-controlled cell has been built for use on the small-angle X-ray scattering instrument D24 at the synchrotron radiation facility LURE. The sample is placed in a quartz capillary sealed in a stainless-steel holder using a vacuum-tight glue. Several O rings provide a vacuum path upstream and downstream from the cell, so that the X-ray beam only meets the capillary walls and the solution under study between the slits and the beam stop, while the sample is maintained under atmospheric pressure. The cell temperature is controlled via a water circulation through a copper sheath in tight contact with the steel holder. The use of this cell results in a marked reduction of the background, as observed in two series of parallel experiments using a conventional cell and this evacuated cell. The decrease ranges from a factor of 2 at s 1 values larger than 0.008 A,-to more than 15 at s --0.00116A -1, where s is the modulus of the scattering vector (s = 2sin 0/A, 20 is the scattering angle and A is the wavelength of the X-rays).

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“…A fragment (residues 1-173) obtained by cyanogen bromide cleavage (Adams et al, 1985) was reported to refold into a native-like structure but to dimerize, thus precluding further characterization. An engineered version of the complete domain (residues 1-184) was characterized as having a "quasi-native" structure (Minard et al, 1989).…”

Section: Discussionmentioning

confidence: 99%

An engineered amino‐terminal domain of yeast phosphoglycerate kinase with native‐like structure

1997

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Previous studies have suggested that the carboxy-terminal peptide (residues 401-415) and interdomain helix (residues 185-199) of yeast phosphoglycerate kinase, a two-domain enzyme, play a role in the folding and stability of the amino-terminal domain (residues 1-184). A deletion mutant has been created in which the carboxy-terminal peptide is attached to the amino-terminal domain (residues 1-184) plus interdomain helix (residues 185-199) through a flexible peptide linker, thus eliminating the carboxy-terminal domain entirely. CD, fluorescence, gel filtration, and NMR experiments indicated that, unlike versions described previously, this isolated N-domain is soluble, monomeric, compactly folded, native-like in structure, and capable of binding the substrate 3-phosphoglycerate with high affinity in a saturable manner. The midpoint of the guanidine-induced unfolding transition was the same as that of the native two-domain protein (C, -0.8 M). The free energy change associated with guanidine-induced unfolding was one-third that of the native enzyme, in agreement with previous studies that evaluated the intrinsic stability of the N-domain and the contribution of domain-domain interactions to the stability of PCK. These observations suggest that the C-terminal peptide and interdomain helix are sufficient for maintaining a native-like fold of the N-domain in the absence of the C-domain.Keywords: autonomously folding domains; isolated domains; phosphoglycerate kinase; protein engineering; protein folding As early as 1973, it was proposed that protein domains represent stable folding units (Wetlaufer, 1973). Since then, a number of isolated domains have been designed, expressed, and characterized (Minard et al

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The folding and mutual interaction of the domains of yeast 3‐phosphoglycerate kinase

Cited by 42 publications

References 23 publications

A comparison of the reactivity and stability of wild type and His388 Gln mutant phosphoglycerate kinase from yeast

A comparison of the reactivity and stability of wild type and His388 Gln mutant phosphoglycerate kinase from yeast

Improved Signal-to-Background Ratio in Small-Angle X-ray Scattering Experiments with Synchrotron Radiation using an Evacuated Cell for Solutions

An engineered amino‐terminal domain of yeast phosphoglycerate kinase with native‐like structure

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