Aldolase Dissociation into Subunits by Reaction with Succinic Anhydride

Hass, Louis F.

doi:10.1021/bi00892a012

Cited by 104 publications

(43 citation statements)

References 33 publications

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“…This amounts to 0.730 ml/g and 0.737 ml/g in the native sunflower [49] and pea proteins, respectively. Hass [50] obtained comparable results for succinylated aldolase.…”

Section: Extent Of Succinylationmentioning

confidence: 80%

Changes of the oligomeric structure of legumin from pea (Pisum sativum L.) after succinylation

Schwenke

Zirwer

Gast

et al. 1990

European Journal of Biochemistry

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The influence of various levels of succinylation on the structure of the legumin from pea seed has been studied by the techniques of sedimentation velocity, viscometry, fluorescence and circular dichroism spectroscopy, as well as dynamic light scattering. The protein dissociates gradually into the 3s subunit forming a 7s intermediate. At a level of 75 -80% succinylation, sudden unfolding of the protein occurs characterized by drastic changes in viscometric and spectroscopic properties. The fluorescence spectra point to the formation of a novel organized structure at a moderate degree of modification before the molecular unfolding takes place.The succinylated subunit was shown to have a sedimentation coefficient of 3.2S, a diffusion coefficient of 5.03 x cm2 . s p l a Stokes' radius of 4.24 nm, a partial specific volume of 0.703 ml/g, an intrinsic viscosity of 0.13 dl/g, a molar mass of 52.2 kDa and a frictional ratio of 1.74.Legumin is one of the main storage proteins in the seeds of pea (Pisum sativum L.). It belongs to the group of the socalled l l S globulins, having sedimentation coefficients between lls and 14S, and molecular masses between 300 kDa and 400 kDa [l]. These proteins possess an oligomeric structure characterized by an arrangement of six subunits (3s components) [2 -41. The molecular mass of pea legumin was reported by different authors to be in the range 330 -410 kDa [l, 5, 61. By means of small-angle X-ray scattering, a value of 359 f 25 kDa was obtained [7]. Disulphide-bonded polypeptide pairs with molecular mass 54 kDa constitute most of the 3s subunit of the legumin [8]. In addition, polypeptide pairs varying in molecular mass over 35-58 kDa have been observed which associate in various ways to give rise to different molecular forms of legumin IS].Due to very similar quaternary structures [9, lo], the 11s plant proteins show an analogous behaviour dissociating into subunits [l 11. Therefore, the decay of oligomeric structure by an increase of negative charge caused by succinylation effects similar changes in the structure of different 11s proteins. Although there are specific differences in the dissociation behaviour of the peanut [12], sunflower seed [13] and rape seed [14] 11s globulins, a step-by-step decay of the native structure, and a marked change in conformation at a critical step of succinylation, were observed in each case.The present paper deals with the effect of succinylation on the oligomeric structure of the 11s globulin (legumin) from pea. Structural changes of the protein were followed using ultracentrifugation, viscometry, circular dichroism and fluorescence spectroscopy. The physicochemical properties of the dissociation products, including molecular mass and shape, were investigated by a combination of hydrodynamic methods, including dynamic light scattering. MATERIALS AND METHODSLegumin was isolated and purified according to Gueguen et al. [15]. Lyophilized preparations containing small amounts of aggregated material were purified additionally by gel filtration on a ...

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“…This amounts to 0.730 ml/g and 0.737 ml/g in the native sunflower [49] and pea proteins, respectively. Hass [50] obtained comparable results for succinylated aldolase.…”

Section: Extent Of Succinylationmentioning

confidence: 80%

Changes of the oligomeric structure of legumin from pea (Pisum sativum L.) after succinylation

Schwenke

Zirwer

Gast

et al. 1990

European Journal of Biochemistry

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“…The LDL was succinylated, using a method based on that of Hass (1964), by addition of succinic anhydride in small increments with stirring to a final molar ratio of succinic anhydride to lysine of 100: 1. The pH was maintained at 8'9 by addition of 0·1 M NaOH.…”

Section: Methodsmentioning

confidence: 99%

Polypeptides From Serum Low-Density Lipoproteins of Pigs (Sus Domesticus)

Walkley¹,

Husbands²

1976

Aust. Jnl. Of Bio. Sci.

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Aust. J. BioI. Sci., 1976, 29, 301-4 Low-density lipoproteins floating between densities 1· 006 and 1 . 063 g cm -3 were isolated by centrifugation of blood serum obtained from 24-h fasted pigs (Sus domesticus). This lipoprotein fraction contained two components with SF 1 . 063 values of 3 . 4 and 2· 3 at 20 D C when examined by analytical ultracentrifugation. Delipidation of the lipoprotein yielded 15 % recovery of soluble protein whereas succinylation of the lipoprotein prior to delipidation gave 95 % recovery of soluble protein. Chromatography on Sephadex Gl00 in 8 M urea of these delipidation products yielded three fractions of different sizes which were present in both native and succinylated apoproteins. These fractions from the succinylated apolipoproteins were further characterized. A polypeptide fraction comprising 70 % of the total protein had an apparent molecular weight of 34000 and contained greater amounts of amino acids with hydrophobic side chains than did the second fraction of apparent molecular weight 22000 which contained 15 % of the protein. The third fraction of apparent molecular weight 12 500 contained 15 % of the protein. IntroductionThe amino acid composition of the serum low-density lipoprotein (LDL) obtained from the domestic pig (Sus domesticus) has been reported by Fidge (1973). However, the apoproteins from human LDL have been shown to contain two different polypeptides (Kane et al. 1970). Pig LDL has also been shown by lanado and Martin (1968) to contain two fractions of different density, LDLI and LDLz. These workers showed that there is heterogeneity in the lipid content of pig LDL. We report here the presence of more than one protein in the LD L of density 1· 006-1 . 063 g em -3 obtained from pig serum.

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“…The results indicated an increase in mobility of the major bands of native and succinylated BSA by 9-6;; and 103% in samples S,, and S,,, respectively. The presence of multiple bands suggests that a variety of products may have formed during the reaction (Hass 1964). The results of relative molecular mass determinations by SDS-PAGE are given in Table 4 and show an increase in mobility of 14 and 1 7 % in samples S,, and S,,, respectively.…”

Section: Polyacrylamide Gel Electrophoresismentioning

confidence: 96%