Abstract
Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing. At their isoelectric points (between pH 4.7 and 5.2) these forms were rather stable. At slightly alkaline pH, each converted into the whole set of isomers. PAGE of the purified enzyme under denaturing or non-denaturing conditions resulted in one protein band. When substrate (amylopectin or pullulan) was included in the gel, the native enzyme as well as any of the individual isomers separated into two (sometimes three) bands ('substrate-induced forms', numbered I-III) with different specific activities, dissociation constants of the enzyme-substrate complexes and activation energies. Each substrate-induced form produced the whole set of seven isomers on isoelectric focusing. The specific activity of the total enzyme reflected the relative proportions of the substrate-induced forms. To some extent the relative proportions, as determined by crossed immunoelectrophoresis, could be shifted in favour of the more or the less active forms by reduction with dithiothreitol, and gentle oxidation respectively. Activation by dithiothreitol did not alter the mode of action of the enzyme but only increased the velocity of substrate degradation and extended its activity into the pH range of the chloroplast. As a consequence of isomer interconversion, microheterogeneity could serve to regulate pullulanase activity in a biochemical manner that shares some features with allosteric regulation.
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Claims 14
Claim · model-written summary
Spinach chloroplast pullulanase separated into at least seven enzymically active isomers.
- Taxon
- Spinacia oleracea
Spinacia oleraceae1 - Organ / stage
- chloroplastse1
- Gene
- pullulanasee1
- Outcome
- individual enzymically active proteinse1
- Value
- at least sevene1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
Claim · model-written summary
The pullulanase isomers were rather stable at their isoelectric points (pH 4.7 to 5.2).
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- these forms were rather stablee2
- Qualifier
- At their isoelectric points (between pH 4.7 and 5.2)e2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“At their isoelectric points (between pH 4.7 and 5.2) these forms were rather stable”
Claim · model-written summary
At slightly alkaline pH each pullulanase isomer converted into the whole set of isomers.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- each converted into the whole set of isomerse3
- Qualifier
- At slightly alkaline pHe3
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e3“At slightly alkaline pH, each converted into the whole set of isomers”
Claim · model-written summary
Purified spinach pullulanase gave one protein band on PAGE under denaturing or non-denaturing conditions.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- protein bande4
- Value
- onee4
- Qualifier
- under denaturing or non-denaturing conditionse4
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e4“PAGE of the purified enzyme under denaturing or non-denaturing conditions resulted in one protein band”
Claim · model-written summary
Substrate in the gel split the native enzyme and its isomers into two (sometimes three) bands.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- substrate (amylopectin or pullulan)e5
- Outcome
- bandse5
- Value
- two (sometimes three)e5
Not reportedComparatorDirection
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e5“When substrate (amylopectin or pullulan) was included in the gel, the native enzyme as well as any of the individual isomers separated into two (sometimes three) bands”
Claim · model-written summary
Substrate-induced forms of spinach pullulanase differed in specific activities, dissociation constants and activation energies.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- specific activitiese6
- Qualifier
- dissociation constants of the enzyme-substrate complexes and activation energiese6
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e6“with different specific activities, dissociation constants of the enzyme-substrate complexes and activation energies”
Claim · model-written summary
Each substrate-induced form produced the whole set of seven isomers on isoelectric focusing.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- whole set of seven isomerse7
- Qualifier
- on isoelectric focusinge7
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e7“Each substrate-induced form produced the whole set of seven isomers on isoelectric focusing”
Claim · model-written summary
The specific activity of the total spinach pullulanase reflected the relative proportions of the substrate-induced forms.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Outcome
- specific activity of the total enzymee2
- Qualifier
- reflected the relative proportions of the substrate-induced formse2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“The specific activity of the total enzyme reflected the relative proportions of the substrate-induced forms”
Claim · model-written summary
Reduction with dithiothreitol shifted the relative proportions of pullulanase forms toward the more active forms.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- reduction with dithiothreitole2
- Outcome
- relative proportionse2
- Direction
- shiftede2
- Qualifier
- To some extente2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“To some extent the relative proportions, as determined by crossed immunoelectrophoresis, could be shifted in favour of the more or the less active forms by reduction with dithiothreitol, and gentle oxidation respectively”
Claim · model-written summary
Gentle oxidation shifted the relative proportions of pullulanase forms toward the less active forms.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- gentle oxidatione2
- Outcome
- relative proportionse2
- Direction
- shiftede2
- Qualifier
- To some extente2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“To some extent the relative proportions, as determined by crossed immunoelectrophoresis, could be shifted in favour of the more or the less active forms by reduction with dithiothreitol, and gentle oxidation respectively”
Claim · model-written summary
Activation by dithiothreitol increased the velocity of spinach pullulanase substrate degradation.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- Activation by dithiothreitole2
- Outcome
- velocity of substrate degradatione2
- Direction
- increasede2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“Activation by dithiothreitol did not alter the mode of action of the enzyme but only increased the velocity of substrate degradation and extended its activity into the pH range of the chloroplast”
Claim · model-written summary
Activation by dithiothreitol did not alter the mode of action of spinach pullulanase.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- Activation by dithiothreitole2
- Outcome
- mode of action of the enzymee2
- Direction
- did not altere2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“Activation by dithiothreitol did not alter the mode of action of the enzyme but only increased the velocity of substrate degradation and extended its activity into the pH range of the chloroplast”
Claim · model-written summary
Activation by dithiothreitol extended the activity of spinach pullulanase into the pH range of the chloroplast.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Treatment
- Activation by dithiothreitole2
- Outcome
- pH rangee2
- Direction
- extendede2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“Activation by dithiothreitol did not alter the mode of action of the enzyme but only increased the velocity of substrate degradation and extended its activity into the pH range of the chloroplast”
Claim · model-written summary
Isomer interconversion may let microheterogeneity regulate spinach pullulanase activity in a biochemical manner.
- Taxon
- Spinacia oleraceaSpinacia oleraceae1
- Gene
- pullulanasee2
- Outcome
- pullulanase activitye2
- Qualifier
- isomer interconversione2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e1“Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing”
e2“As a consequence of isomer interconversion, microheterogeneity could serve to regulate pullulanase activity in a biochemical manner”
The same questions in other papers
1,4-dithiothreitol · physiology: enzyme activity · Spinacia oleracea (spinach)Results agree
3 papers in the same direction as this paper
- no effect
- no effect
- no effect
Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.