Molecular weight and amino acid composition of five-times-crystallized phosphoglucose isomerase from rabbit skeletal muscle

Pon, Ning G.; Schnackerz, Klaus D.; Blackburn, Michael N.; Chatterjee, Gaurav; Noltmann, Ernst A.

doi:10.1021/bi00809a005

Cited by 54 publications

(18 citation statements)

References 38 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The appearance of additional electrophoretic species from GPI I on rechromatography or other manipulative procedures, and the retardation of these changes by a sulfhydryl reagent, parallels the results reported for rabbit muscle GPI by Pon et al (11).…”

supporting

confidence: 86%

“…These are stable protein species that are not interconvertible under physiological conditions (9). In contrast to this, Pon et al (11) have shown that multiple forms of rabbit muscle GPI separated by column chromatography do not remain as single species during rechromatography under the same conditions, although the addition of dithiothreitol prevents previously separated single peaks from separating further. These forms of rabbit muscle isomerases are apparently different forms of the same enzyme protein and can be designated as "pseudoisozvmes.…”

mentioning

confidence: 95%

“…At pH 6.0, 4 mm citrate strongly inhibits all isomerases. The inhibition is about 45% for I and 60% for both Ia 11 (64 ,ug) was preincubated at 60 C with 100 umoles of glycyl-glycine buffer, pH 8.0, for the times indicated. After preincubation, glucose-6-P (3 ,tmoles) was added to start the reaction.…”

mentioning

confidence: 99%

See 2 more Smart Citations

Multiple Forms of Glucosephosphate Isomerase in Maize

Salamini¹,

Tsai²,

Nelson³

1972

Plant Physiol.

View full text Add to dashboard Cite

Three apparently different glucosephosphate isomerases are found in the developing seeds of maize (Zea mays L.). Glu umn chromatography of developing endosperm preparations. Activity under the Ia peak can be enhanced with a concomitant attenuation of activity under the I peak by certain purification procedures. I and II are, however, quite distinct noninterconvertible enzymes that may have different physiological functions. A third enzyme, GPI IIl, is detectable only in the embryo. This enzyme elutes from a DEAE-cellulose column in the same fractions as does GPl lI from the endosperm but is distinguishable from II on the basis of electrophoretic mobility and the stimulatory effect of ATP in the reaction mixtLire.These results contrast with those from pea and green gram (13,16), where highly purified enzymes have been prepared, but no heterogeneity has been reported. GPI from yeast has been crystallized (7). Recently, Nakagawa and Noltmann (8) fractionated the crystallized enzyme into three isozymic forms by elution with a very shallow phosphate gradient from a DEAE-cellulose column. These are stable protein species that are not interconvertible under physiological conditions (9). In contrast to this, Pon et al. (11) have shown that multiple forms of rabbit muscle GPI separated by column chromatography do not remain as single species during rechromatography under the same conditions, although the addition of dithiothreitol prevents previously separated single peaks from separating further. These forms of rabbit muscle isomerases are apparently different forms of the same enzyme protein and can be designated as "pseudoisozvmes." We demonstrate here that both situations are found in developing maize seeds. There are multiple forms that are separable by column chromatography and distinguishable by various tests. Additionally, GPI I can generate a number of electrophoretically separable pseudoisozymes under certain preparative regimens.Since we are interested in the regulation of starch synthesis in maize, glucosephosphate isomerase activity in the maize seed has been investigated. The results reveal that three peaks of activity (I, Ia, II) can be detected after DEAE5-cellulose col-

show abstract

supporting

confidence: 86%

mentioning

confidence: 95%

See 1 more Smart Citation

Multiple Forms of Glucosephosphate Isomerase in Maize

Salamini¹,

Tsai²,

Nelson³

1972

Plant Physiol.

View full text Add to dashboard Cite

show abstract

“…Yeast and rabbit phosphoglucose isomerases differ considerably more. The molecular weight of the yeast enzyme may be somewhat smaller than the rabbit enzyme (i.e., 120000 [20] versus 132000 [21]), and there is disagreement whether the yeast enzyme is composed of two [22] or of four [23] subunits. Nevertheless, yeast monomers did not appear to be capable of forming active hybrids with the rabbit enzyme.…”

Section: Discussionmentioning

confidence: 99%

Matrix-Bound Phosphoglucose Isomerase. Formation and Properties of Monomers and Hybrids

Bruch¹,

Schnackerz²,

Gracy³

1976

Eur J Biochem

Self Cite

View full text Add to dashboard Cite

Dimeric phosphoglucose isomerase from rabbit muscle was immobilized by reaction with cyanogen-bromide-activated Sepharose 4B. The catalytic parameters and stability properties of the free and matrix-bound isomerases were essentially identical. Total monomerisation of the matrix-bound enzyme was achieved with 8 M urea as determined by a study in which one subunit was labelled with i~do['~C]acetate and the other with the 3H-labelled reagent. Although matrixbound monomers were devoid of isomerase activity, they were still capable of binding the substrate. Matrix-bound monomers exhibited the ability to redimerize with soluble isomerase subunits from either rabbit or human yielding catalytically active dimers. Yeast isomerase monomers, in contrast, did not yield active hybrids with the rabbit monomers. Furthermore, soluble subunits, which had been inactivated with pyridoxal 5'-phosphate were also capable of hybridizing with and inducing catalytic activity in the matrix-bound monomers. These studies indicate the prerequisite of dimer formation for catalytic activity but the independent action of the catalytic centers of the dimer.Dissociation of phosphoglucose isomerase into subunits has been achieved in 3 mM sodium dodecylsulfate, 6 M guanidine hydrochloride and 8 M urea [l -31. The demonstration of the dimeric structure of the enzyme, identical polypeptide chains [4] and of two substrate binding sites [5] does not exclude the functional non-equivalence of the two active sites.

show abstract

“…The pellet was dissolved in a small volume of Hepes buffer and then dialyzed against several changes of buffer (0.01 M Hepes/0.0074 M KOH/0.1 M KC1, pH 8.0) to remove ammonium sulfate. The protein concentration was determined spectrophotometrically by using e280 p.i = 1.32 ml/mg-cm (12). Where necessary, the enzyme was concentrated by using Amicon Centricon 10 centrifugal ultrafiltration devices.…”

Section: Methodsmentioning

confidence: 99%

Phosphoglucose isomerase: a ketol isomerase with aldol C2-epimerase activity.

Seeholzer

1993

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

View full text Add to dashboard Cite

With 13C NMR, phosphoglucose isomerase (PGI; D-glucose-6-phosphate ketol-isomerase, EC 5.3.1.9) is shown to produce mannose 6-phosphate (M6P) slowly from a much more rapidly catalyzed equilibrium between glucose 6-phosphate (G6P) and fructose 6-phosphate (F6P (4,6,20,21). PGI was found to catalyze a second kind of reaction when the interconversions of the a-and ,B-pyranose anomers of G6P (7) and the a-and l3-furanose anomers of F6P (8) by PGI were first demonstrated. These findings have more recently been confirmed by using 31P NMR (9) and 13C NMR (10) spectroscopic techniques. The anomerase activity requires ring cleavage, subsequent torsional rotation about the C1-C2 bond for G6P and C2-C3 bond for F6P, and finally cyclization to the opposite face ofthe carbonyl plane. Studies with epoxide-inactivated PGI (8) suggest that both anomers of each substrate react at the same active site responsible for catalyzing the isomerase reaction. PGI was also shown to mutarotate the anomers of mannose 6-phosphate (M6P) at a rate similar to its effect on G6P (11).Reported here are experiments that demonstrate and characterize yet a third enzymatic reaction catalyzed by PGI: G6P C2-epimerization. This reaction results in the production of MGP from G6P and F6P. A mechanism is proposed that shows the relationship between the isomerase, anomerase, and epimerase activities of PGI. MATERIALS AND METHODSRabbit muscle PGI was obtained from Sigma as a suspension in ammonium sulfate. The suspension was centrifuged and decanted. The pellet was dissolved in a small volume of Hepes buffer and then dialyzed against several changes of buffer (0.01 M Hepes/0.0074 M KOH/0.1 M KC1, pH 8.0) to remove ammonium sulfate. The protein concentration was determined spectrophotometrically by using e280 p.i = 1.32 ml/mg-cm (12). Where necessary, the enzyme was concentrated by using Amicon Centricon 10 centrifugal ultrafiltration devices. PGI activity was typically 580 units/mg when assayed spectrophotometrically in 0.1 M Hepes/0.1 M KC1, pH 8/8 mM F6P/10 units of G6P dehydrogenase (G6PDH)/ 0.5 mM NADP, 25°C.G6P, F6P, and M6P were obtained as their sodium salts from Sigma. G6P was assayed spectrophotometrically (340 nm) with NADP (0.3 mM) and G6PDH (bakers' yeast, Sigma) in a buffer containing 0.1 M triethanolamine, 0.1 M KCl, 0.01 M MgCl2 (pH 7.6). F6P and M6P were assayed in the same cuvette by sequentially adding PGI and then phosphomannose isomerase (PMI, yeast, Sigma), recording the A30 endpoint after each addition.Abbreviations: G6P, glucose 6-phosphate; F6P, fructose 6-phosphate; M6P, mannose 6-phosphate; PGI, phosphoglucose isomerase; PMI, phosphomannose isomerase; G6PDH, G6P dehydrogenase; kat, catalytic rate constant.

show abstract

Molecular weight and amino acid composition of five-times-crystallized phosphoglucose isomerase from rabbit skeletal muscle

Cited by 54 publications

References 38 publications

Multiple Forms of Glucosephosphate Isomerase in Maize

Multiple Forms of Glucosephosphate Isomerase in Maize

Matrix-Bound Phosphoglucose Isomerase. Formation and Properties of Monomers and Hybrids

Phosphoglucose isomerase: a ketol isomerase with aldol C2-epimerase activity.

Contact Info

Product

Resources

About