Key points are not available for this paper at this time.
UDP-l-rhamnose is required for the biosynthesis of cell wall rhamnogalacturonan-I, rhamnogalacturonan-II, and natural compounds in plants. It has been suggested that the RHM2/MUM4 gene is involved in conversion of UDP-d-glucose to UDP-l-rhamnose on the basis of its effect on rhamnogalacturonan-I-directed development in Arabidopsis thaliana. RHM2/MUM4-related genes, RHM1 and RHM3, can be found in the A. thaliana genome. Here we present direct evidence that all three RHM proteins have UDP-d-glucose 4,6-dehydratase, UDP-4-keto-6-deoxy-d-glucose 3,5-epimerase, and UDP-4-keto-l-rhamnose 4-keto-reductase activities in the cytoplasm when expressed in the yeast Saccharomyces cerevisiae. Functional domain analysis revealed that the N-terminal region of RHM2 (RHM2-N; amino acids 1–370) has the first activity and the C-terminal region of RHM2 (RHM2-C; amino acids 371–667) has the two following activities. This suggests that RHM2 converts UDP-d-glucose to UDP-l-rhamnose via an UDP-4-keto-6-deoxy-d-glucose intermediate. Site-directed mutagenesis of RHM2 revealed that mucilage defects in MUM4-1 and MUM4-2 mutant seeds of A. thaliana are caused by abolishment of RHM2 enzymatic activity in the mutant strains and furthermore, that the GXXGXX(G/A) and YXXXK motifs are important for enzymatic activity. Moreover, a kinetic analysis of purified His6-tagged RHM2-N protein revealed 5.9-fold higher affinity of RHM2 for UDP-d-glucose than for dTDP-d-glucose, the preferred substrate for dTDP-d-glucose 4,6-dehydratase from bacteria. RHM2-N activity is strongly inhibited by UDP-l-rhamnose, UDP-d-xylose, and UDP but not by other sugar nucleotides, suggesting that RHM2 maintains cytoplasmic levels of UDP-d-glucose and UDP-l-rhamnose via feedback inhibition by UDP-l-rhamnose and UDP-d-xylose. UDP-l-rhamnose is required for the biosynthesis of cell wall rhamnogalacturonan-I, rhamnogalacturonan-II, and natural compounds in plants. It has been suggested that the RHM2/MUM4 gene is involved in conversion of UDP-d-glucose to UDP-l-rhamnose on the basis of its effect on rhamnogalacturonan-I-directed development in Arabidopsis thaliana. RHM2/MUM4-related genes, RHM1 and RHM3, can be found in the A. thaliana genome. Here we present direct evidence that all three RHM proteins have UDP-d-glucose 4,6-dehydratase, UDP-4-keto-6-deoxy-d-glucose 3,5-epimerase, and UDP-4-keto-l-rhamnose 4-keto-reductase activities in the cytoplasm when expressed in the yeast Saccharomyces cerevisiae. Functional domain analysis revealed that the N-terminal region of RHM2 (RHM2-N; amino acids 1–370) has the first activity and the C-terminal region of RHM2 (RHM2-C; amino acids 371–667) has the two following activities. This suggests that RHM2 converts UDP-d-glucose to UDP-l-rhamnose via an UDP-4-keto-6-deoxy-d-glucose intermediate. Site-directed mutagenesis of RHM2 revealed that mucilage defects in MUM4-1 and MUM4-2 mutant seeds of A. thaliana are caused by abolishment of RHM2 enzymatic activity in the mutant strains and furthermore, that the GXXGXX(G/A) and YXXXK motifs are important for enzymatic activity. Moreover, a kinetic analysis of purified His6-tagged RHM2-N protein revealed 5.9-fold higher affinity of RHM2 for UDP-d-glucose than for dTDP-d-glucose, the preferred substrate for dTDP-d-glucose 4,6-dehydratase from bacteria. RHM2-N activity is strongly inhibited by UDP-l-rhamnose, UDP-d-xylose, and UDP but not by other sugar nucleotides, suggesting that RHM2 maintains cytoplasmic levels of UDP-d-glucose and UDP-l-rhamnose via feedback inhibition by UDP-l-rhamnose and UDP-d-xylose. In plants, UDP-l-rhamnose (UDP-Rha) 2The abbreviations used are: UDP-Rha, UDP-l-rhamnose; dTDP, deoxythymidine 5′-diphosphate; HPLC, high performance liquid chromatography; ESI/MS, electrospray ionization-mass spectrometry; COSY, correlated spectroscopy; TOCSY, total correlation spectroscopy; NOESY, nuclear Overhauser effect spectroscopy; RG-I, rhamnogalacturonan-I; RG-II, rhamnogalacturonan-II; dTDP-4K6DG, dTDP-4-keto-6-deoxy-d-glucose; dTDP-4KR, dTDP-4-keto-l-rhamnose; UDP-Xyl, UDP-d-xylose; MOPS, 4-morpholinepropanesulfonic acid; MES, 4-morpholineethanesulfonic acid; DTT, dithiothreitol; Ni-NTA, nickel-nitrilotriacetic acid. is required for biosynthesis of the primary cell wall components rhamnogalacturonan-I (RG-I) and rhamnogalacturonan-II (RG-II), and various l-rhamnose-containing natural compounds (flavonoids, terpenoids, and saponins) (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google Scholar, 2Ikan R. Naturally Occurring Glycosides. 1999; (Wiley, Chichester, UK)Google Scholar). Three major pectic polysaccharides (homogalacturonan, RG-I, and RG-II) are present in the primary cell walls of plants (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google Scholar). RG-I is a polymer of more than 100 individual α1,4-linked disaccharide units that are themselves composed of l-rhamnose and d-galacturonic acid (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google Scholar). RG-II is a structurally complex polysaccharide that exists in primary walls as a dimer covalently cross-linked by a borate diester bond (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google Scholar, 3O'Neill M.A. Warrenfeltz D. Kates K. Pellerin P. Doco T. Darvill A.G. Albersheim P. J. Biol. Chem. 1996; 271: 22923-22930Abstract Full Text Full Text PDF PubMed Scopus (428) Google Scholar). These pectic polysaccharides play a major role in the development and growth of all vascular plants (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google Scholar). In bacteria, cell surface polysaccharides also contain l-rhamnose and are essential for survival and interaction between bacteria (4Dong C. Beis K. Giraud M.F. Blankenfeldt W. Allard S. Major L.L. Kerr I.D. Whitfield C. Naismith J.H. Biochem. Soc. Trans. 2003; 31: 532-536Crossref PubMed Google Scholar, 5Jiang X.M. Neal B. Santiago F. Lee S.J. Romana L.K. Reeves P.R. Mol. Microbiol. 1991; 5: 695-713Crossref PubMed Scopus (276) Google Scholar). The rhamnose-containing polysaccharides of bacteria are synthesized from dTDP-rhamnose (dTDP-Rha) (4Dong C. Beis K. Giraud M.F. Blankenfeldt W. Allard S. Major L.L. Kerr I.D. Whitfield C. Naismith J.H. Biochem. Soc. Trans. 2003; 31: 532-536Crossref PubMed Google Scholar, 5Jiang X.M. Neal B. Santiago F. Lee S.J. Romana L.K. Reeves P.R. Mol. Microbiol. 1991; 5: 695-713Crossref PubMed Scopus (276) Google Scholar). In the last decade, much effort has been directed at studying the dTDP-Rha synthetic pathway (e.g. gene structures, enzymatic properties, and their functions in cell wall synthesis) (4Dong C. Beis K. Giraud M.F. Blankenfeldt W. Allard S. Major L.L. Kerr I.D. Whitfield C. Naismith J.H. Biochem. Soc. Trans. 2003; 31: 532-536Crossref PubMed Google Scholar). A dTDP-Rha biosynthetic gene cluster consisting of the rmlB (or rfbB), rmlC (or rfbC), and rmlD (or rfbD) genes is responsible for biosynthesis of dTDP-Rha from dTDP-glucose (dTDP-Glc) in bacteria, Escherichia coli, Mycobacterium tuberculosis, and Salmonella enterica serovar Typhimurium (Fig. 1) (4Dong C. Beis K. Giraud M.F. Blankenfeldt W. Allard S. Major L.L. Kerr I.D. Whitfield C. Naismith J.H. Biochem. Soc. Trans. 2003; 31: 532-536Crossref PubMed Google Scholar, 5Jiang X.M. Neal B. Santiago F. Lee S.J. Romana L.K. Reeves P.R. Mol. Microbiol. 1991; 5: 695-713Crossref PubMed Scopus (276) Google Scholar). The rmlB, rmlC, and rmlD genes encode dTDP-Glc 4,6-dehydratase (EC 4.2.1.46), dTDP-4-keto-6-deoxy-d-glucose (dTDP-4K6DG) 3,5-epimerase (EC 5.1.3.13), and dTDP-4-keto-l-rhamnose (dTDP-4KR) 4-keto-reductase (EC 1.1.1.133), respectively (5Jiang X.M. Neal B. Santiago F. Lee S.J. Romana L.K. Reeves P.R. Mol. Microbiol. 1991; 5: 695-713Crossref PubMed Scopus (276) Google Scholar). In plants, UDP-Rha is synthesized from UDP-d-glucose (UDP-Glc) via an analogous enzymatic pathway (6Kamsteeg J. Van Brederode J. Van Nigtevecht G. FEBS Lett. 1978; 91: 281-284Crossref PubMed Scopus (23) Google Scholar) (Fig. 1). It is known that UDP-Rha is synthesized from UDP-Glc by UDP-Glc 4,6-dehydratase (EC 4.2.1.76), UDP-4-keto-6-deoxy-d-glucose (UDP-4K6DG) 3,5-epimerase, and UDP-4-keto-l-rhamnose (UDP-4KR) 4-keto-reductase (6Kamsteeg J. Van Brederode J. Van Nigtevecht G. FEBS Lett. 1978; 91: 281-284Crossref PubMed Scopus (23) Google Scholar). RHM1, RHM2/MUM4, RHM3, and UER1 proteins of Arabidopsis thaliana are putative plant orthologues of the dTDP-Rha biosynthetic enzymes in bacteria (Figs. 2A and 3A). It has been that the UER1 protein Biol. PubMed Scopus Google also G. C. PubMed Scopus Google has 3,5-epimerase and 4-keto-reductase from The N-terminal and C-terminal of RHM proteins are to and the basis of and Mol. Biol. 2001; PubMed Scopus Google Scholar) that RHM1, RHM2/MUM4, and proteins are putative sugar enzymes in A. thaliana. It has also been that in the mucilage of seeds of MUM4-2 and RG-I is found at levels and is of a than in the mucilage of seeds B. PubMed Scopus Google Scholar, PubMed Scopus Google suggesting that RHM2 is involved in the of evidence for the of enzymes has been the other a of as an of a in the gene that a A. thaliana cell protein A. B. B. C. PubMed Scopus Google Scholar). RHM1, B. and W. D. A. B. B. C. PubMed Scopus Google Scholar) found that the activity of RHM1 in conversion of UDP-Glc to UDP-Rha, the as and to the of a of RHM1 has not been domain analysis of RHM2 of and UER1 proteins to a RHM2 amino N-terminal region of RHM2 amino C-terminal region of RHM2 amino UER1 amino analysis of and UER1 The of UER1 an from of yeast on used as a for RHM2-N as a and and UER1 proteins at for His6-tagged RHM2-N His6-tagged His6-tagged RHM2-N and His6-tagged His6-tagged UER1 His6-tagged RHM2-N and His6-tagged UER1 in analysis of UDP-Rha activity. activities as from of yeast used as a of for His6-tagged RHM2 His6-tagged RHM2-N His6-tagged His6-tagged RHM2-N and His6-tagged UER1 His6-tagged RHM2-N and UER1 from three are In we expressed RHM1, and in yeast and found that the RHM1, RHM2/MUM4, and proteins are enzymes UDP-Glc 4,6-dehydratase, 3,5-epimerase, and 4-keto-reductase activities. of enzymatic and of protein revealed that the N-terminal region of RHM2/MUM4 amino UDP-Glc 4,6-dehydratase and the C-terminal region of RHM2/MUM4 amino 3,5-epimerase and that the genes are responsible for the of UDP-Rha from UDP-Glc in plants. In RHM2-N activity is strongly inhibited by UDP-Rha, and UDP but not by other sugar and suggesting that RHM2/MUM4 maintains cytoplasmic levels of UDP-Glc and UDP-Rha via feedback inhibition by UDP-Rha and we the of on the of in plants. and strains used in as A. J. P. PubMed Scopus Google and in a synthetic yeast amino acids and an amino acids F. 1991; PubMed Scopus Google Scholar). growth in liquid by of of in a The at at for Saccharomyces used D. A. PubMed Scopus Google Scholar). of acid acid and UDP-Rha, and The from the following and from UDP-Rha synthesized a cytoplasmic from yeast RHM2/MUM4 and purified by a of His6-tagged RHM1, RHM2/MUM4, and for of RHM1 RHM2/MUM4 and as The A. thaliana RHM1 and RHM2/MUM4 genes by A. thaliana as a for the and and and by a P. K. J. PubMed Scopus Google Scholar, A. J. T. A. K. J. P. T. K. A. K. 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The at a of The for UDP-Glc and UDP-Rha and the as a at the from to by a to the by Lee Lee Lee K. PubMed Scopus Google Scholar) a the A at a of for and at a of for The for and UDP-Rha and of by enzymatic from UDP-Glc of RHM2 and RHM2-N and The by electrospray on an in the for as by the by nuclear on a a at and The UDP-Rha and purified in of The of correlation and Overhauser effect of to and to all The to and in the domain in the A total of for TOCSY, NOESY, for for all by the to the the and amino acid used to the of RHM2-N from His6-tagged RHM2-N protein purified from yeast cell at the cell yeast in of at for in of A from from and from and by at for and of to the The for The by at for and for of DTT, from and The of A The in A of purified His6-tagged RHM2-N The purified by the acid protein as a of of UDP-Glc 4,6-dehydratase activities as UDP-Glc and purified His6-tagged RHM2-N in a total of 100 The UDP-Glc 4,6-dehydratase in the of various The purified His6-tagged RHM2-N at for The by for The by an by Three of of and on of His6-tagged for activity as the for His6-tagged the purified protein at a of and for and the by for The for His6-tagged RHM2-N as Three of and and and and The purified His6-tagged RHM2-N used for at for and the by for activity by the of the via an by Three of of 4,6-dehydratase activity by the of UDP-Glc and purified His6-tagged RHM2-N in a total of 100 in the are in the by of UDP-Glc of the purified His6-tagged RHM2-N used in for of UDP-Glc for and by for activity by the of the via an by and sugar levels dTDP-Glc UDP-Glc as a between and of dTDP-Glc and The and the the of three and of RHM1, RHM2/MUM4, and A. thaliana RHM1, RHM2/MUM4, and genes by from an A. thaliana The RHM1, RHM2/MUM4, and proteins to orthologues also in other plant and has a putative of RHM2/MUM4 the of the gene is The N-terminal of RHM1, RHM2/MUM4, and are at the amino acid to the a dTDP-Glc 4,6-dehydratase of The of RHM1, RHM2/MUM4, and have to is a 4-keto-reductase of and a high of to A. thaliana has 3,5-epimerase and 4-keto-reductase suggesting that the N-terminal and C-terminal of RHM proteins are we the N-terminal region of RHM2 acids 1–370) as RHM2-N and the C-terminal region acids 371–667) as RHM2-N and to the sugar of a putative P. Mol. Biochem. 1978; PubMed Scopus Google Scholar) and a D. PubMed Google Scholar). The that RHM1, and are cytoplasmic proteins that not the functions of RHM we to the yeast S. to the as is known that proteins from plants can be expressed and in yeast to T. T. PubMed Google Scholar) and of as the cell the RHM1, RHM2/MUM4, and genes the of the S. and RHM2/MUM4 expressed in the S. we to a major and putative an (Fig. the of RHM2/MUM4 protein is the is to be a of the RHM2/MUM4 protein the RHM2/MUM4 gene expressed in various of S. cerevisiae. The of RHM2/MUM4 protein the for the protein in the and strains (Fig. and but not in the strains (Fig. and The and other to the in the that the strains we is the for and of RHM proteins (Fig. we used the as a for protein in the enzymatic of RHM we an of and protein that a present on the expressed be from the cytoplasmic of cell of RHM1, RHM2/MUM4, and genes in in of a major protein to (Fig. used as a for proteins (Fig. and the of RHM2/MUM4 than that of RHM1 and suggesting that the RHM2/MUM4 protein is more than RHM1 in yeast (Fig. Functional of RHM1, and we enzymatic activities of the the cytoplasmic of yeast RHM1, RHM2/MUM4, as a of UDP-Rha synthetic activity UDP-Glc as a substrate and and as The cytoplasmic of RHM genes conversion of UDP-Glc to UDP-Rha (Fig. 1) but in cytoplasmic from RHM1, RHM2/MUM4, a as at (Fig. suggesting that A is an enzymatic as that A to UDP-Rha, we the for the substrate (UDP-Glc) and A from the RHM2 and their by and in the purified sugar from the RHM2/MUM4 gene (Fig. The to that of UDP-Glc from a (Fig. and the is the of UDP-Rha, suggesting that A is the we of A a from RHM2 (Fig. and the by by COSY, TOCSY, and various a of for for A. in for A are in The and of UDP-Rha are also in by and analysis of A are in The of UDP-Rha on is in for A in the and G. C. PubMed Scopus Google Scholar) of UDP-Rha 1). In analysis revealed that and of the on the of a (Fig. the of A as UDP-Rha, strongly suggesting that RHM1, RHM2/MUM4, and proteins can UDP-Glc to and of UDP-Rha synthesized from UDP-Glc by RHM2 by analysis by analysis in a of Functional in the RHM2/MUM4 we the of the and C-terminal of The of that RHM2-N and have enzymatic as RHM2-N is to is to and plant UER1 G. C. PubMed Scopus Google Scholar, Mol. Biol. 2001; PubMed Scopus Google Scholar, B. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). the of domain of we for the and UER1 genes the S. and (Fig. 3A). The and the of proteins in the cytoplasmic of cell by analysis as (Fig. The RHM2-N in of a protein (Fig. and the and UER1 in of proteins (Fig. and RHM2-N and RHM2-N and proteins of and in (Fig. and the enzymatic activities of RHM2-N and the cytoplasmic of UER1 genes as a of UDP-Rha synthetic activity UDP-Glc as a substrate and and as The cytoplasmic of and UER1 conversion of UDP-Glc (Fig. and In in a as The and a from to (Fig. suggesting that is an enzymatic of RHM2-N as It is that of via the interaction of the in the we in the In RHM2-N and RHM2-N and conversion of to UDP-Rha (Fig. and that RHM2-N and are and that has the enzymatic activity as UER1 (Fig. and the of we the substrate (UDP-Glc) from a and from a RHM2-N and their by and in the purified sugar from the RHM2-N (Fig. The has the as UDP-Glc from the (Fig. and the is the of suggesting that the is (Fig. the of the we of from the RHM2-N and the via the for analysis of UDP-Rha (Fig. and a of used to a for are in The and of are in on the basis of UDP-Rha The and are in In the the of a and a of (Fig. and at and in the from UDP-Rha, to of a at the This strongly suggests that the of the is The also the of and as and that is in the of of These also the that the sugar is evidence that RHM2-N a UDP-Glc 4,6-dehydratase that converts UDP-Glc to the that a 3,5-epimerase and a to and of synthesized from UDP-Glc by RHM2-N by analysis by analysis in a of in the RHM2/MUM4 amino acid can the enzymatic activities of the RHM2/MUM4 The of in the RHM2/MUM4 protein are in PubMed Scopus Google Scholar) that MUM4-1 and MUM4-2 contain a amino acid of to and to These a in mucilage and cytoplasmic PubMed Scopus Google Scholar). has not been are caused by a of RHM2/MUM4 protein by a in RHM2/MUM4 enzymatic activity. of RHM2/MUM4 UDP-Rha we first for of and mutant of of RHM2-N and revealed that to the have a for P. Mol. Biochem. 1978; PubMed Scopus Google Scholar) in their N-terminal and have two of YXXXK a as the in proteins D. PubMed Google Scholar) (Fig. the of amino acid we also of RHM2/MUM4, and (Fig. mutant expressed in and The as a for The of mutant protein present in the than that of RHM2 (Fig. and and the and the for in RHM2-N and also (Fig. and suggesting that the mutant proteins are than RHM2 (Fig. the mutant we and in protein levels (Fig. and we UDP-Rha activity of the mutant proteins of as a of and UDP-Glc 4,6-dehydratase activity (Fig. evidence that the MUM4-1 and MUM4-2 in A. thaliana PubMed Scopus Google Scholar) are to abolishment of UDP-Rha activity. the not activity but also of the and also UDP-Glc 4,6-dehydratase that and are essential for conversion of UDP-Glc to (Fig. of the mutant protein than that of the suggesting that is also important for UDP-Glc 4,6-dehydratase activity (Fig. and are the RHM2-N their on the UDP-Glc 4,6-dehydratase activity of when 3,5-epimerase and 4-keto-reductase activity cytoplasmic from strains mutant the levels of activities from of RHM2/MUM4 not the other and are to UDP-Glc to (Fig. and are in the is to that not UDP-Glc 4,6-dehydratase activity. evidence that the RHM2-N and of RHM2/MUM4 can in the RHM2 C-terminal region on activity. The activities (Fig. the a effect on activity and caused a in activity as RHM2 (Fig. on we that of the in domain and enzymatic activity of that domain (Fig. of and and a in the of the protein (Fig. and These that of a is important not for activity but also for the of RHM2 and furthermore, that and have on the activity of the the that the is RHM1, RHM2/MUM4, and the not the activity of The is RHM1, RHM2/MUM4, and but not in UER1 and that has been by in to a by G. C. PubMed Scopus Google Scholar) that the UER1 protein has 3,5-epimerase and 4-keto-reductase activities (Fig. evidence that is not important for RHM2/MUM4 activity. of the RHM2-N we on the of the RHM2-N protein of RHM2-N a effect on yeast cell growth and protein (Fig. defects can be by of the gene of the UER1 gene (Fig. and we expressed RHM2-N a and a in and purified RHM2-N protein we to of the protein from a A (Fig. S. enterica and enzymatic activity UDP-Glc than of the activity dTDP-Glc the B. Whitfield C. P. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google that proteins dTDP-Glc to and not UDP-Glc to B. Whitfield C. P. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). the and substrate of 4,6-dehydratase of RHM2 are we the and kinetic for we first activity a of The of activity in In the activity than that for at the activity when the the of activity the to (Fig. activity also at The for activity and the that the is in the to (Fig. substrate and of we the and of UDP-Glc and dTDP-Glc in the of the in the of His6-tagged RHM2-N the UDP-Glc 4,6-dehydratase in the of UDP-Glc and dTDP-Glc are between and 100 and the also and the of UDP-Glc and dTDP-Glc and respectively Moreover, the for UDP-Glc and dTDP-Glc and respectively These are of suggesting that RHM2 UDP-Glc to dTDP-Glc as a of RHM2-N protein for UDP-Glc and activity of in a of the activity of RHM2-N is inhibited by various and The activity of the purified His6-tagged RHM2-N protein inhibited by UDP-Rha, UDP-Xyl, and UDP-Rha is a of UDP-Glc conversion by RHM2 enzymatic and when UDP-Rha used as an activity inhibited of the for the also to activity to a of the for the UDP-Rha is more than of RHM2-N activity not by other we of These to that RHM2-N activity is by the of UDP-Rha and in the cytoplasmic of UDP and at also inhibited the activity of RHM2-N but the of not This suggests that UDP and can to the and RHM2-N compounds are than is than not the activity of effect of compounds on UDP-Glc 4,6-dehydratase in a In we used a yeast to evidence that RHM1, RHM2/MUM4, and encode proteins that can UDP-Rha from This is the first of genes that in the UDP-Rha synthetic in RHM proteins yeast and and that RHM1, and as a protein three enzymatic activities that UDP-Glc to and are involved in and of other that levels of activities are in than in 1991; Scholar). has been the RHM proteins are to the and by various of RHM proteins the to by cell UDP-Rha is by to cell wall of RG-I, RG-II (1Ridley B.L. O'Neill M.A. Mohnen D. Phytochemistry. 2001; 57: 929-967Crossref PubMed Scopus (1530) Google and various l-rhamnose-containing natural compounds in plants R. Naturally Occurring Glycosides. 1999; (Wiley, Chichester, UK)Google suggesting that UDP-Rha is essential to plant is known the responsible as UDP-Rha is a sugar that is not analysis of revealed the of genes in the of A. thaliana J. P. B.L. PubMed Scopus Google Scholar). It be to genes that encode proteins activity from genes UDP-Rha can be yeast be to UDP-Rha that can be used to the of putative and to the of the of plant cell wall polysaccharides l-rhamnose in also evidence that the N-terminal and C-terminal of RHM2/MUM4 are The N-terminal domain of RHM2/MUM4 (RHM2-N; amino acids 1–370) as a UDP-Glc 4,6-dehydratase and the C-terminal domain of RHM2/MUM4 (RHM2-C; amino acids 371–667) as a consisting of 3,5-epimerase and 4-keto-reductase activities. It is to the two in plant RHM the enzymatic by the are to conversion of to by and as is synthesized from by 4,6-dehydratase, 3,5-epimerase, and 4-keto-reductase S. A. PubMed Scopus Google Scholar, J. PubMed Google Scholar). genes are present as in A. RHM2/MUM4 is a consisting of domain and that and proteins to enzymatic activity K. 2003; PubMed Scopus Google Scholar). is that the two of RHM2 protein be to a growth and cell in RHM2-N yeast This is to a effect caused by of as of UER1 RHM2-N the defects not It that to the as the and of domain analysis of RHM2/MUM4 and analysis of the revealed that is responsible for of and G. C. PubMed Scopus Google Scholar). The of in an expressed Biol. PubMed Scopus Google Scholar, Mol. 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PubMed Scopus Google Scholar, R. 1996; PubMed Scopus Google Scholar, PubMed Scopus Google Scholar, S. PubMed Scopus Google Scholar, J. A. Mol. Biol. 57: PubMed Scopus Google Scholar). also its activities UDP-Glc and PubMed Scopus Google Scholar). a feedback inhibition conversion of UDP-Glc to UDP-Xyl, and in plants, suggesting that a role to the is important for plant to a of not its synthetic pathway but also the of UDP-Rha from be required to the of inhibition and UDP-Rha and UDP-Xyl, not RHM2-N the that is synthesized by of R. 2003; PubMed Scopus Google Scholar). The enzymes that to are suggesting on the of the R. 2003; PubMed Scopus Google Scholar). is in the and not in the be for by It be to RHM2-N is inhibited by but the that is not at present to is in the cell wall than is l-rhamnose R. Lee J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google suggesting that is and in the the effect of on RHM2-N activity not be a we the that UDP-Rha as via inhibition of It has been that a UDP-Glc of various from at the of the in higher plants T. D. S. S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google a pathway for UDP-Rha has not been are required we the that of the in plant for and and and for are to and for
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