Molecular weight and amino acid composition of purified spinach beet phenolase

Vaughan, Peter F. T.; Eason, R.W.; Paton, J Y; Ritchie, G. A. F.

doi:10.1016/0031-9422(75)80349-9

Cited by 34 publications

(12 citation statements)

References 15 publications

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“…Results ofthe various runs are shown in Figure SA for the enzyme stain and Figure PEAR POLYPHENOL OXIDASE DISCUSSION Detergent extraction, using Triton X-100, increased the amount of PPO activity that could be extracted from D'Anjou pears. Several other researchers have reported similar results with detergents (I 1, 13,26). The increase in enzyme activity has been attributed to an activation due to the detergent (1 1), and to the disruption of chloroplast membranes to release more enzyme (21).…”

Section: Resultssupporting

confidence: 53%

“…However, when the extract was applied onto a hydrophobic column, all of the pigment remained on top of the column, while the enzyme could be eluted in several clear and colorless fractions, indicating that the enzyme was not in a tight complex with the Chl. Vaughan et al (26) recommended the use of cetyltrimethylammonium bromide (Cetavlon), a cationic detergent, in the extraction of PPO from spinach beet since it did not liberate the Chl pigments. The effect of temperature on the elution of PPO from the Phenyl Sepharose, DEAE, and hydroxylapatite columns was an interesting observation.…”

Section: Resultsmentioning

confidence: 99%

“…Few reports have been published on PPO preparations purified to apparent homogeneity from plants (1,4,12,26), and only three researchers used fruits (9,16,24). Rivas and Whitaker (24) reported a purification procedure for Bartlett pears that yielded two enzyme fractions that were homogeneous by electrophoresis with a final 20-and 37-fold increase in specific activity; however, the extracts were tanned.…”

Section: Resultsmentioning

confidence: 99%

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Purification of d'Anjou Pear (Pyrus communis L.) Polyphenol Oxidase

Wissemann¹,

Montgomery²

1985

Plant Physiol.

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Polyphenol oxidase (PPO) was extensively purified to homogeneity from d'Anjou pear (Pyrus communis L.) by extraction in the presence of the phenolic binder AG 2-X8 andTriton X-100. Chlorophyll pigment was removed by chromatography resulting in a clear, colorless enzyme extract. Purification of pear PPO was achieved after chromatography on Phenyl Sepharose CL4B, DEAE-cellulose, and hydroxylapatite columns. Only after the columns were run at room temperature rather than at 4°C were sharp peaks and good resolution obtained. Reproducibility of the entire scheme was excellent with chromatography on the hydrophobic resin as a key to successful purification. Three separate fractions of pear PPO were homogeneous when stained for protein with the silver stain after polyacryhamide slab gel electrophoresis and corresponded to relative mobilities of OAI, OA3, and 0.73. The effect of dimethylsulfoxide on enzyme activity was investigated and found to iucrease significantly the activity of purified pear PPO over the control.Oxidative browning catalyzed by PPO4 (monophenol, dihydroxyphenylalanine:oxygen oxidoreductase; EC 1.14.18.1) that occurs when fresh fruits and vegetables are damaged is of economic concern to most fruit and vegetable processors. Information on the molecular structure and catalytic site would help to define the mode of action of PPO and methods to control its undesirable effects. Isolation and purification of PPO from contaminating proteins and phenolics is essential for these studies. Unfortunately, the purification of PPO has typically been a difficult task, and the number of PPO preparations that have been purified to homogeneity from higher plants and particularly from fruits is very limited (9,16,24

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Section: Resultssupporting

confidence: 53%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Purification of d'Anjou Pear (Pyrus communis L.) Polyphenol Oxidase

Wissemann¹,

Montgomery²

1985

Plant Physiol.

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“…1 and 3). No product corresponding to the widely reported 45-kD polypeptide (Vaughan et al, 1975;Vaughn and Duke, 1984;Flurkey, 1985Flurkey, , 1986Lanker et al, 1987;Vaughn et al, 1988;Steffens et al, 1990) has been observed either as a translation product of Hy-19 or following import and processing by plastids. Our observations are in accordance with accumulating evidence on the structure of PPO genes in higher plants.…”

Section: Thylakoid Lumen By the Conventional Mechanismsmentioning

confidence: 91%

“…The active form of PPO was frequently reported to be about 45 kD (Vaughan et al, 1975;Lanker et al, 1987), but other forms, e.g. 59 and 65 to 68 kD were also reported Ganesa et al, 1992;Shahar et al, 1992).…”

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confidence: 99%