Characterization of two forms of the multifunctional protein acting in fatty acid β-oxidation

Behrends, Wilke; Engeland, Kurt; Kindl, Helmut

doi:10.1016/0003-9861(88)90624-8

Cited by 39 publications

(15 citation statements)

References 13 publications

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“…The difference in the ratio of MFP and RNA-bound MFP (compare upper and lower panels) in the soluble and the pellet fractions of the RNA-microtubule-MFP assay suggests that MFP on its own may have a higher binding affinity for microtubules than does the MFP-RNA complex, possibly due to steric hindrance caused by the bound RNAs. Taken together these results and other observations that plant MFPs appear to exist as a monomer 2 (50) indicate that there are distinct RNA-and microtubule-binding sites on the MFP polypeptide.…”

Section: Figsupporting

confidence: 73%

Identification of a Rice RNA- and Microtubule-binding Protein as the Multifunctional Protein, a Peroxisomal Enzyme Involved in the β-Oxidation of Fatty Acids

Chuong¹,

Mullen²,

Muench³

2002

Journal of Biological Chemistry

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The control of subcellular mRNA localization and translation is often mediated by protein factors that are directly or indirectly associated with the cytoskeleton. We report the identification and characterization of a rice seed protein that possesses both RNA and microtubule binding activities. In vitro UV cross-linking assays indicated that this protein binds to all mRNA sequences tested, although there was evidence for preferential binding to RNAs that contained A-C nucleotide sequence motifs. The protein was purified to homogeneity using a two-step procedure, and amino acid sequencing identified it as the multifunctional protein (MFP), a peroxisomal enzyme known to possess a number of activities involved in the ␤-oxidation of fatty acids. The recombinant version of this rice MFP binds to RNA in UV cross-linking and gel mobility shift experiments, co-sediments specifically with microtubules, and possesses at least two enzymatic activities involved in peroxisomal fatty acid ␤-oxidation. Taken together these data suggest that MFP has an important role in mRNA physiology in the cytoplasm, perhaps in regulating the localization or translation of mRNAs through an interaction with microtubules, in addition to its peroxisomal function.

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Section: Figsupporting

confidence: 73%

Identification of a Rice RNA- and Microtubule-binding Protein as the Multifunctional Protein, a Peroxisomal Enzyme Involved in the β-Oxidation of Fatty Acids

Chuong¹,

Mullen²,

Muench³

2002

Journal of Biological Chemistry

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“…This theory is supported by the altered fatty acid composition of the aim1 mutant. A total of four MFPs have been purified and characterized from cucumber (Behrends et al, 1988;Kindl, 1995a, 1995b), although only one has been cloned and sequenced (Preisig-Müller et al, 1994). Three of these proteins have been isolated from cotyledons (MFPI to MFPIII), whereas the fourth (MFPIV) was isolated from leaf tissue.…”

Section: Biochemical Context Of the Aim1 Mutationmentioning

confidence: 99%

A Defect in β-Oxidation Causes Abnormal Inflorescence Development in Arabidopsis

1999

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The abnormal inflorescence meristem1 ( aim1 ) mutation affects inflorescence and floral development in Arabidopsis. After the transition to reproductive growth, the aim1 inflorescence meristem becomes disorganized, producing abnormal floral meristems and resulting in plants with severely reduced fertility. The derived amino acid sequence of AIM1 shows extensive similarity to the cucumber multifunctional protein involved in ␤ -oxidation of fatty acids, which possesses L -3-hydroxyacyl-CoA hydrolyase, L -3-hydroxyacyl-dehydrogenase, D -3-hydroxyacyl-CoA epimerase, and ⌬ 3 , ⌬ 2 -enoyl-CoA isomerase activities. A defect in ␤ -oxidation has been confirmed by demonstrating the resistance of the aim1 mutant to 2,4-diphenoxybutyric acid, which is converted to the herbicide 2,4-D by the ␤ -oxidation pathway. In addition, the loss of AIM1 alters the fatty acid composition of the mature adult plant.

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“…MFP I and MFP I1 were purified from cotyledons of 4-day-old dark-grown cucumber (Cucumis sativus) seedlings according to [22). Large fragments, prepared by limited proteolysis, were isolated by FPLC using the cation-exchange resin Mono S HR 5/5 (Pharmacia) or hydroxyapatite HAPSuperformance (Merck).…”

Section: Isolation Of Multifunctional Proteins and Their Peptide Fragmentioning

confidence: 99%

Evidence for Domain Structures of the Trifunctional Protein and the Tetrafunctional Protein Acting in Glyoxysomal Fatty Acid β‐Oxidation

Gühnemann-Schäfer

Engeland

Linder

et al. 1994

European Journal of Biochemistry

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In plant glyoxysomes, an enzyme activity responsible for a particular step in the fatty acid , ! loxidation is located on more than one protein species. Various monofunctional enzymes and two forms of a multifunctional protein are involved in the degradation of cis-unsaturated fatty acids. A',A*-Enoyl-CoA isomerase activity, previously found to be located on a monofunctional dimeric protein, is attributable to one form of the monomeric multifunctional protein (MFP). The presence or absence of isomerase activity allows us to differentiate between the tetrafunctional 76.5-kDa isoform (MFP 11) and the trifunctional 74-kDa isoform (MFP I) in cucumber (Cucumis sativus) cotyledons. Both MFP I and MFP I1 exhibited blocked N-terminal structures. MFP I and MFP I1 are distinguishable from each other by their susceptibility to limited proteolysis. A series of examples is presented describing the preparation of enzymically active proteolytic fragments. We demonstrate that both forms of the monomeric MFP are composed of domains separable from each other without loss of activity. By fragmentation of MFP I and subsequent chromatography, a 60-kDa peptide was purified retaining hydratase and epimerase activity but lacking dehydrogenase activity. In addition, a highly positively charged fragment was observed carrying solely dehydrogenase activity. From MFP 11, a 36-kDa fragment with hydratase activity was characterized. An enzymically inactive 46-kDa fragment was prepared from MFP I1 and sequenced at its unblocked N-terminus.Comprehensive studies of the enzymes from glyoxysomes and peroxisomes have provided evidence for a close phylogenetic relationship [ 1 -41. For fatty acid P-oxidation, all forms of microbodies possess a multifunctional protein (MFP) as marker protein [4-71. Studies indicate that this protein represents a fusion product of two, three or four single enzyme peptides connected in a linear sequence [5,8, 91. This leads to the assumption that several domains as compact globular structures form the tertiary structures of MFP. Portions of the polypeptide chain may fold to stable tertiary structures, independent of the presence of other parts of the polypeptide chain. Domains of a large protein like MFP may be connected by short flexible regions free of enzymic activity. If such flexible regions, unlike the compact globular structure bearing active sites, are easily accessible for proteases, we expected that domains with single active centers can be cleaved off, purified and analyzed. It is suggestive that at least one exposed hinge region exists in the MFP, namely between the 3-hydroxyacyl-CoA dehydrogenase region and a region bearing, among others, the 2-enoyl-CoA hydratase activity. In this context, we consider the two forms of MFP with different size and different number of functions as model systems for demonstrating the domain structure of proteins.Correspondence to H. Kindl, FB Chemie, Philipps-Universitat, D-35032 Marburg, GermanyEnzymes. L-3-Hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35) ; d3,dZ-enoyl-CoA iso...

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Characterization of two forms of the multifunctional protein acting in fatty acid β-oxidation

Cited by 39 publications

References 13 publications

Identification of a Rice RNA- and Microtubule-binding Protein as the Multifunctional Protein, a Peroxisomal Enzyme Involved in the β-Oxidation of Fatty Acids

Identification of a Rice RNA- and Microtubule-binding Protein as the Multifunctional Protein, a Peroxisomal Enzyme Involved in the β-Oxidation of Fatty Acids

A Defect in β-Oxidation Causes Abnormal Inflorescence Development in Arabidopsis

Evidence for Domain Structures of the Trifunctional Protein and the Tetrafunctional Protein Acting in Glyoxysomal Fatty Acid β‐Oxidation

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