Succinylation yeast

The only disadvantage of the succinylation procedure (which is practical and amenable to conventional cell disruption processes) is that the final product is a succinylated protein. Succinyl groups cannot be removed from the succinylated proteins under mild conditions. This could be a problem if succinylated yeast protein was a major source of dietary proteins. Therefore we explored the feasibility of using reversible modifying reagents (citraconic anhydride and maleic anhydride) to separate proteins from NAs and subsequently remove the modifying groups under mild acidic conditions. 

Incubation of succinylated yeast homogenate at 55°C (pH 6.0). to activate endogenous ribonuclease and hydrolyze the contaminant RNA resulted in the significant reduction of the extensive proteolysis normally observed (Table VI). Furthermore, the coagulation and precipitation of protein that normally occurs during this step was also eliminated. 

Table VII. The Influence of pH of Protein Precipitation on the Content (%) of Nucleic Acid in Nonsuccinylated and Succinylated Yeast Protein...

Table VIII. Composition of Yeast Cells,Succinylated Yeast Protein Concentrate and Succinylated Yeast Protein Isolate (expressed as g/lOOg)...

Succinylation reduced the isoelectric point of yeast proteins from pH A.5 to 4.0 and markedly improved their solubility in pH range 4.5 to 6. Heat denatured yeast proteins were facilely solubilized following succinylation (74). Succinylated yeast proteins were very stable to heat above pH 5, and remained soluble at temperatures above 80°C. As the degree of succinylation was increased the rate of precipitation of the derivatized protein increased in the neighborhood of the isoelectric point and much larger protein floes were obtained facilitiating their recovery (74). 

Table IX. Emulsifying Activity Index Values Protein of Succinylated Yeast...

The in vitro digestibility of succinylated yeast proteins by a-chymotrypsin and trypsin was compared (Fig. 9). In both instances hydrolysis of both derivatized and unmodified yeast protein was quite similar, though the a-chymotrypsin hydrolyzed the succinylated protein more rapidly. 

Succinylated yeast protein has not only an increased solubility in the pH range of 4-6, but is also more heat stable above pH 5. It has better emulsifying properties, surpassing many other proteins (Table 1.37), and has increased whippa-bility. 

This succinyl group avoids the formation of the pyranosidic ring and acts as a spacer to conjugate the mole-cule to resins or biopolymers. The succinylated analog of sucrose 41 (R= HO2CCH2CH2CO-) is not sweet, and it is unable to inhibit invertase. It is however an inhibitor of a-glucosidase from baker s yeast. 

Pantothenic acid is a component of coenzyme A, which functions in the transfer of acyl groups (Figure 28.17). Coenzyme A contains a thiol group that carries acyl compounds as activated thiol esters. Examples of such structures are succinyl CoA, fatty acyl CoA, and acetyl CoA. Pantothenic acid is also a component of fatty acid synthase (see p. 182). Eggs, liver, and yeast are the most important sources of pan tothenic acid, although the vitamin is widely distributed. Pantothenic acid deficiency is not well characterized in humans, and no RDA has been established. 

Separation of "Proteins from the Nucleoprotein Complex in Yeast by Succinylation Procedure... 

Isolation of Proteins with a Reduced Nucleic Acid Level. The procedure is virtually identical to that described for succinylation of yeast proteins (87). In a typical experiment proteins, together with NA, were extracted from the disrupted yeast cells at pH 8.5-9.0 and centrifuged at 15,000 rpm for 30 min at 5°C. Citraconic anhydride then was added in small increments to the supernatant with constant stirring while the pH was maintained between 8.0-8.5 by adding 3.5IV NaOH. After the stabilization of the pH, the pH of the solution was decreased to 4.2 to precipitate the proteins. Protein then was separated by centrifugation, dissolved in water (pH adjusted 8.5), dialyzed extensively against water (pH 8.5) at 5°C, and lyophilized. 

Table VI. The Effect of Succinylation on Proteolysis of Yeast Proteins by Endogenous Proteases During Incubation (pH 6.0, 55°C) for RNA Reduction...

Thus, succinylation prevented the aggregation and coagulation of yeast proteins normally observed during RNA hydrolysis with endogenous ribonuclease and markedly increased the recovery of intact, soluble protein. Lindbloom (69) reported that up to 70% of yeast protein may be degraded during incubation to reduce RNA (pH 6,... 

Figure 4. The increased quantity of protein extracted from homogenized yeast cells at pH 8.5 following succinylation...

Figure 5. The effects of succinylation during extraction of yeast protein on the endogenous ribonuclease activity (pH 6.0, 55°C)...

Figure 7. Outline of the succinylation procedure for the isolation of yeast protein concentrate (72% protein) or yeast protein isolate (92% protein) with low...

Chemical modification of yeast protein has received limited attention though as described above it has potential as a method for facilitating recovery of yeast protein. Current studies are concerned with determination of the functional properties of proteins succinylated during the extraction. The composition of yeast proteins prepared by different methods is shown (Table 8). Noteworthy is the protein and nucleic acid concentration in the yeast isolate which differed from the concentrate in that cell wall material was removed by centrifugation. 

The absorption spectra of three yeast protein preparations prepared by different procedures were compared (Fig. 8). The presence of nucleic acid which has a X maximum at 260 nm tend to shift the absorption spectrum of yeast protein to lower wavelengths. The ratio of absorption at 280 to 260 nm is indicative of NA contamination in protein samples a ratio of more than one indicates pure protein devoid of nucleic acid whereas a ratio of 0.65 indicates approximately 30% contamination with NA. The yeast protein extracted with alkali and directly acid precipitated showed a X max at 260, a 280/260 ratio of 0.67 and contained 28%, NA determined chemically. Protein extracted in alkali, adjusted to pH 6 and incubated at 55°C for 3-5 hours, to reduce NA with endogenous ribonuclease, had a X max at 260, a 280/260 ratio of 0.8 and a NA content of 3.3% while yeast protein prepared by the succinylation procedure and precipitated at pH 4.5 showed a X max at 275 nm, a 280/260 ratio of 1.0 and nucleic acid content of 1.8. 

The emulsifying capacity of the yeast proteins was progressively improved with the extent of succinylation (Table IX) as measured by the turbidimetric technique (89). The modified yeast proteins had excellent emulsifying activities compared to several other common proteins. McElwain et al. (88) observed that succinylation of yeast protein increased emulsion viscosity but decreased emulsion stability. 

Legend crude protein prepared by precipitation of an alkali extract at pH 4.0 ( ) yeast protein obtained following activation of endogenous ribonuclease (82) (O), and yeast protein prepared by the succinylation procedure (O). 

Figure 9. Rate of hydrolysis of succinylated and nonsuccinylated yeast proteins by a-chymotrypsin, (O) unsuccinylated and ( ) succinylated, ana trypsin, (A) unsuccinylated and (A) succinylated...

To achieve success as protein ingredients for food formulation and fabrication, novel proteins should possess a range of functional properties. Frequently during extraction, refining and drying, plant and yeast proteins, intended for food uses, become denatured or altered and subsequently display poor functional properties which render them of limited use. Chemical modification provides a feasible method for improving the functional properties of plant and yeast proteins and potentially may make it possible to tailor proteins with very specific functional properties. In this review the information on modified plant proteins is reviewed and the use of succinylation for the recovery of yeast proteins with low nucleic acid is described. 

In animals, yeasts and purple photosynthetic bacteria ALA 15 is made from glycine 13 and succinyl CoA 14 by the action of a single enzyme, ALA synthase. 

Figure 5.32 Biosynthetic pathways to 5-aminolevulinic acid (5-ALA) from glutamate (higher plants) and from glycine and succinyl-coenzyme A (animals, fungi, yeast, and bacteria).

The carbon skeleton of homocysteine is derived from the corresponding hydroxy amino acid, homoserine. The hydroxyl of homoserine is acylated with either a succinyl (bacteria) or acetyl (yeast, fungi, plants) group derived from the corresponding coenzyme A derivative. The O-acyl substituent is then displaced by the thiol group of cysteine producing a mixed thioether, cystathionine. This in turn undergoes a pyridoxal phosphate dependent -elimination to homocysteine, pyruvate and... 

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