The Experts below are selected from a list of 279 Experts worldwide ranked by ideXlab platform
Masaki Okuda - One of the best experts on this subject based on the ideXlab platform.
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Glutelin subtype-dependent protein localization in rice grain evidenced by immunodetection analyses
Plant Molecular Biology, 2019Co-Authors: Kei Takahashi, Hiromi Kohno, Tomomichi Kanabayashi, Masaki OkudaAbstract:Key message GluA and GluB-4/5 Glutelin subfamilies are mainly localized to outer region of the endosperm, particularly in its ventral side, in rice grain, but GluC is localized to throughout the endosperm. Abstract The major seed storage protein in rice ( Oryza sativa ) is Glutelin, which forms a vacuole-derived protein body type-II. Glutelins are encoded by multiple genes, and generally comprise four protein subfamilies, namely, GluA, GluB, GluC, and GluD: however, the localization pattern of Glutelin in rice grains remains obscure. In this study, we investigated the localization pattern of five subtypes of the Glutelin protein in rice grains using Glutelin-subtype specific antibodies. Immunoblot analysis against sequentially polished rice flour fractions from three crop years and seven japonica rice varieties revealed that GluA was strongly localized in the outer region of the endosperm, including the subaleurone layer, whereas GluC was distributed throughout the endosperm. Among the Glutelin subtypes, GluA was mostly found in the outer region of the rice grain, followed by GluB-4/5, GluB-1, GluD, and GluC. Immunofluorescence labeling microscopy analysis using immature rice seeds clearly revealed that the localization pattern of GluC and GluD was completely different from that of GluA and GluB. Expression levels of all Glutelins, particularly GluA, GluB-1, and GluB-4/5, were stronger on the ventral than dorsal side in rice grains. These results provide strong and consistent evidence that Glutelins localize to the rice grain in a subfamily-dependent manner.
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Glutelin subtype-dependent protein localization in rice grain evidenced by immunodetection analyses.
Plant molecular biology, 2019Co-Authors: Kei Takahashi, Hiromi Kohno, Tomomichi Kanabayashi, Masaki OkudaAbstract:GluA and GluB-4/5 Glutelin subfamilies are mainly localized to outer region of the endosperm, particularly in its ventral side, in rice grain, but GluC is localized to throughout the endosperm. The major seed storage protein in rice (Oryza sativa) is Glutelin, which forms a vacuole-derived protein body type-II. Glutelins are encoded by multiple genes, and generally comprise four protein subfamilies, namely, GluA, GluB, GluC, and GluD: however, the localization pattern of Glutelin in rice grains remains obscure. In this study, we investigated the localization pattern of five subtypes of the Glutelin protein in rice grains using Glutelin-subtype specific antibodies. Immunoblot analysis against sequentially polished rice flour fractions from three crop years and seven japonica rice varieties revealed that GluA was strongly localized in the outer region of the endosperm, including the subaleurone layer, whereas GluC was distributed throughout the endosperm. Among the Glutelin subtypes, GluA was mostly found in the outer region of the rice grain, followed by GluB-4/5, GluB-1, GluD, and GluC. Immunofluorescence labeling microscopy analysis using immature rice seeds clearly revealed that the localization pattern of GluC and GluD was completely different from that of GluA and GluB. Expression levels of all Glutelins, particularly GluA, GluB-1, and GluB-4/5, were stronger on the ventral than dorsal side in rice grains. These results provide strong and consistent evidence that Glutelins localize to the rice grain in a subfamily-dependent manner.
Fumio Takaiwa - One of the best experts on this subject based on the ideXlab platform.
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reducing rice seed storage protein accumulation leads to changes in nutrient quality and storage organelle formation
Plant Physiology, 2010Co-Authors: Taiji Kawakatsu, Sakiko Hirose, Hiroshi Yasuda, Fumio TakaiwaAbstract:Rice (Oryza sativa) seed storage proteins (SSPs) are synthesized and deposited in storage organelles in the endosperm during seed maturation as a nitrogen source for germinating seedlings. We have generated Glutelin, globulin, and prolamin knockdown lines and have examined their effects on seed quality. A reduction of one or a few SSP(s) was compensated for by increases in other SSPs at both the mRNA and protein levels. Especially, reduction of Glutelins or sulfur-rich 10-kD prolamin levels was preferentially compensated by sulfur-poor or other sulfur-rich prolamins, respectively, indicating that sulfur-containing amino acids are involved in regulating SSP composition. Furthermore, a reduction in the levels of 13-kD prolamin resulted in enhancement of the total lysine content by 56% when compared with the wild type. This observation can be mainly accounted for by the increase in lysine-rich proteins. Although reducing the level of Glutelins slightly decreased protein storage vacuoles (PSVs), the simultaneous reduction of Glutelin and globulin levels altered the inner structure of PSVs, implicating globulin in framing PSV formation. Knock down of 13-kD prolamins not only reduced the size of endoplasmic reticulum-derived protein bodies (PBs) but also altered the rugged peripheral structure. In contrast, PBs became slightly smaller or unchanged by severe suppression of 10- or 16-kD prolamins, respectively, indicating that individual prolamins have distinct functions in the formation of PBs. Extreme increases or decreases in sulfur-poor prolamins resulted in the production of small PBs, suggesting that the ratio of individual prolamins is crucial for proper aggregation and folding of prolamins.
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Characterization of a new rice Glutelin gene GluD-1 expressed in the starchy endosperm
Journal of experimental botany, 2008Co-Authors: Taiji Kawakatsu, Masayuki P. Yamamoto, Sakiko Hirose, Masahiro Yano, Fumio TakaiwaAbstract:A new Glutelin gene, designated GluD-1, has been discovered by comparing the seed storage proteins from 48 japonica and indica rice cultivars on SDS-PAGE gels. Evidence that GluD-1 is a member of the Glutelin family was provided by Western blots using anti-Glutelin antiserum and by mapping the gene to the chromosomal Glutelin gene cluster. The limited GluD-1 size polymorphism among the rice varieties is due to amino acid substitutions rather than to post-transcriptional modification. GluD-1 is maximally expressed in the starchy endosperm starting at 5 d after flowering (DAF) and increasing through 30 DAF, a major difference from the other Glutelins which are primarily expressed in the subaleurone from 10-16 DAF. Only about 0.2 kb of the GluD-1 promoter was sufficient to confer inner starchy endosperm-specific expression. The 0.2 kb truncated GluD-1 promoter contains a bifactorial endosperm box consisting of a truncated GCN4 motif (TGA(G/C)TCA) and AAAG Prolamin box (P box), and ACGT and AACA motifs as cis-regulatory elements. Gel retardation assays and trans-activation experiments indicated that the truncated GCN4 and P box are specifically recognized by RISBZ1 b-ZIP and RPBF Dof activators in vitro, respectively, and are synergistically transactivated, indicating that combinatorial interactions of these motifs are involved in essential endosperm-specific regulation. Furthermore, deviation from the cognate GCN4 motif alters tissue-specific expression in the inner starchy endosperm to include other endosperm tissues.
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Accumulation of Soybean Glycinin and Its Assembly with the Glutelins in Rice
Plant physiology, 1999Co-Authors: Tomoyuki Katsube, Nobuyuki Kurisaka, Masahiro Ogawa, Nobuyuki Maruyama, Reiko Ohtsuka, Shigeru Utsumi, Fumio TakaiwaAbstract:Saline-soluble glycinins and insoluble Glutelins are the major storage proteins in soybean (Glycine max) and rice (Oryza sativa), respectively. In spite of their differences in solubility properties, both proteins are members of the 11S globulin gene family based on their similarities in primary sequences and processing of the coded protein. Wild-type and methionine-modified glycinin coding sequences were expressed in transgenic rice plants under the control of the rice Glutelin GluB-1 promoter. Glycinins were specifically synthesized in the endosperm tissue and co-localized with Glutelins in type II protein bodies. They assembled into 7S and 11S species, similar to what was observed in developing soybean seeds. This pattern was quite different from that displayed by the rice Glutelins in untransformed plants, in which processed subunits sedimenting at 2S were apparent. In glycinin-expressing transgenic plants, however, Glutelins were observed sedimenting at 7S and 11S with lesser amounts in the 2S region. A portion of the glycinins was also found associated in the insoluble Glutelin fraction. Renaturation experiments suggested that the hybrid glycinin-Glutelin oligomers were formed through specific interactions. Overall, these results indicate that despite significant differences in the assembly of soybean glycinin and rice Glutelin, both proteins can assemble with each other to form soluble hexameric oligomers or insoluble aggregates.
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Sequence of three members and expression of a new major subfamily of Glutelin genes from rice
Plant molecular biology, 1991Co-Authors: Fumio Takaiwa, Kiyoharu Oono, David Wing, Akira KatoAbstract:Three members have been isolated of an additional Glutelin gene subfamily, named subfamily B, consisting of about five members per haploid rice genome. Restriction fragment length polymorphism analysis showed major differences between Japonica and Indica lines, indicating the divergence of the subfamily since the split between the two varieties. While corresponding exons of the subfamily B showed 80 to 88% nucleotide sequence homology, those exons were only 60–65% homologous to those of the Glutelin A subfamily [15, 19, 24], distinguishing them from the subfamily A. Intron position and derived polypeptide structure, in addition to the nucleotide sequence, confirm the subfamily B members as Glutelins. Analysis of RNA from seeds of different stages of development showed that the subfamily B members were expressed at the same time as those of subfamily A, demonstrating coordinated regulation of the two subfamilies.
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Localization of Glutelin gene in rice chromosome by in situ hybridization
The Japanese Journal of Genetics, 1991Co-Authors: Hiroaki Suzuki, Fumio Takaiwa, Yuzo Futsuhara, Nori KurataAbstract:The location of storage protein Glutelin gene on a chromosome was determined by in situ hybridization made between Glutelin cDNA clone and mitotic chromosomes in rice. After hybridization, the location of 3H-labelled Glutelin gene was analyzed on chromosomes of twenty one cells. It was found that silver grains were mainly concentrated on the short arm of chromosome 2 (K2 chromosome), demonstrating that the gene responsible for storage protein Glutelin was located on this chromosome.
Kei Takahashi - One of the best experts on this subject based on the ideXlab platform.
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Glutelin subtype-dependent protein localization in rice grain evidenced by immunodetection analyses
Plant Molecular Biology, 2019Co-Authors: Kei Takahashi, Hiromi Kohno, Tomomichi Kanabayashi, Masaki OkudaAbstract:Key message GluA and GluB-4/5 Glutelin subfamilies are mainly localized to outer region of the endosperm, particularly in its ventral side, in rice grain, but GluC is localized to throughout the endosperm. Abstract The major seed storage protein in rice ( Oryza sativa ) is Glutelin, which forms a vacuole-derived protein body type-II. Glutelins are encoded by multiple genes, and generally comprise four protein subfamilies, namely, GluA, GluB, GluC, and GluD: however, the localization pattern of Glutelin in rice grains remains obscure. In this study, we investigated the localization pattern of five subtypes of the Glutelin protein in rice grains using Glutelin-subtype specific antibodies. Immunoblot analysis against sequentially polished rice flour fractions from three crop years and seven japonica rice varieties revealed that GluA was strongly localized in the outer region of the endosperm, including the subaleurone layer, whereas GluC was distributed throughout the endosperm. Among the Glutelin subtypes, GluA was mostly found in the outer region of the rice grain, followed by GluB-4/5, GluB-1, GluD, and GluC. Immunofluorescence labeling microscopy analysis using immature rice seeds clearly revealed that the localization pattern of GluC and GluD was completely different from that of GluA and GluB. Expression levels of all Glutelins, particularly GluA, GluB-1, and GluB-4/5, were stronger on the ventral than dorsal side in rice grains. These results provide strong and consistent evidence that Glutelins localize to the rice grain in a subfamily-dependent manner.
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Glutelin subtype-dependent protein localization in rice grain evidenced by immunodetection analyses.
Plant molecular biology, 2019Co-Authors: Kei Takahashi, Hiromi Kohno, Tomomichi Kanabayashi, Masaki OkudaAbstract:GluA and GluB-4/5 Glutelin subfamilies are mainly localized to outer region of the endosperm, particularly in its ventral side, in rice grain, but GluC is localized to throughout the endosperm. The major seed storage protein in rice (Oryza sativa) is Glutelin, which forms a vacuole-derived protein body type-II. Glutelins are encoded by multiple genes, and generally comprise four protein subfamilies, namely, GluA, GluB, GluC, and GluD: however, the localization pattern of Glutelin in rice grains remains obscure. In this study, we investigated the localization pattern of five subtypes of the Glutelin protein in rice grains using Glutelin-subtype specific antibodies. Immunoblot analysis against sequentially polished rice flour fractions from three crop years and seven japonica rice varieties revealed that GluA was strongly localized in the outer region of the endosperm, including the subaleurone layer, whereas GluC was distributed throughout the endosperm. Among the Glutelin subtypes, GluA was mostly found in the outer region of the rice grain, followed by GluB-4/5, GluB-1, GluD, and GluC. Immunofluorescence labeling microscopy analysis using immature rice seeds clearly revealed that the localization pattern of GluC and GluD was completely different from that of GluA and GluB. Expression levels of all Glutelins, particularly GluA, GluB-1, and GluB-4/5, were stronger on the ventral than dorsal side in rice grains. These results provide strong and consistent evidence that Glutelins localize to the rice grain in a subfamily-dependent manner.
Hikaru Satoh - One of the best experts on this subject based on the ideXlab platform.
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rna targeting to a specific er sub domain is required for efficient transport and packaging of α globulins to the protein storage vacuole in developing rice endosperm
Plant Journal, 2012Co-Authors: Haruhiko Washida, Masahiro Ogawa, Toshihiro Kumamaru, Aya Sugino, Kelly A Doroshenk, Mio Satohcruz, Ai Nagamine, Tomoyuki Katsubetanaka, Hikaru SatohAbstract:Summary Studies focusing on the targeting of RNAs that encode rice storage proteins, prolamines and Glutelins to specific sub-domains of the endoplasmic reticulum (ER), as well as mis-localization studies of other storage protein RNAs, indicate a close relationship between the ER site of RNA translation and the final site of protein deposition in the endomembrane system in developing rice endosperm. In addition to prolamine and Glutelin, rice accumulates smaller amounts of α-globulins, which are deposited together with Glutelin in the protein storage vacuole (PSV). In situ RT-PCR analysis revealed that α-globulin RNAs are not distributed to the cisternal ER as expected for a PSV-localized protein, but instead are targeted to the protein body-ER (PB-ER) by a regulated process requiring cis-sorting sequences. Sequence alignments with putative maize δ-zein cis-localization elements identified several candidate regulatory sequences that may be responsible for PB-ER targeting. Immunocytochemical analysis confirmed the presence of α-globulin on the periphery of the prolamine protein bodies and packaging in Golgi-associated dense vesicles, as well as deposition and storage within peripheral regions of the PSV. Mis-targeting of α-globulin RNAs to the cisternal ER dramatically alters the spatial arrangement of α-globulin and Glutelin within the PSV, with the accompanying presence of numerous small α-globulin particles in the cytoplasm. These results indicate that α-globulin RNA targeting to the PB-ER sub-domain is essential for efficient transport of α-globulins to the PSV and its spatial arrangement in the PSV. Such RNA localization prevents potential deleterious protein–protein interactions, in addition to performing a role in protein targeting.
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PAKISTAN RICE GENETIC RESOURCES-III: SDS-PAGE DIVERSITY PROFILE OF GlutelinS (SEED STORAGE PROTEIN)
Pakistan Journal of Botany, 2010Co-Authors: Sadar Uddin Siddiqui, Toshihiro Kumamaru, Hikaru SatohAbstract:Rice grain quality characters pertaining to seed storage proteins profile for Glutelin was evaluated for diversity within Pakistan local rice genetic resources using SDS-PAGE. Materials consisted of 475 accessions collected from 3-rice cultivation zones and other parts of the country. A wide variation was found in the Glutelin fraction of rice protein at 57kD pro-Glutelin and 40kD Glutelin acidic subunit bands 3 and 4. The enriched Glutelin variation at 57kD may be used in the development of improved protein cultivars with respect to quality and quantity.
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Genetic Variation of Glutelin Acidic Subunit Polypeptides in Bangladesh Rice Genetic Resources
Genetic Resources and Crop Evolution, 2005Co-Authors: Md. Sarwar Jahan, Yuji Uemura, Toshihiro Kumamaru, Abdul Hamid, Hikaru SatohAbstract:Bangladesh rice genetic resources collected from six distinct regions of the country were examined to obtain the diversity in Glutelin acidic subunit polypeptides. Seed Glutelins from 576 Bangladesh rice cultivars representing seven ecotypes were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), isoelectric focusing (IEF) electrophoresis and two-dimensional electrophoresis (2-DE: SDS-PAGE/IEF) analyses. Glutelin acidic subunit was separated into four bands, α -1, α -2, α -3 and α -4, and the variation in each of bands was detected by SDS-PAGE analysis. A higher molecular size component of α -1( α -1′) was identified in the cultivars tested. In case of α -2, α -2H with high molecular mass, α -2L with low molecular mass and α -2H/L were detected. α -3 band variation showed α -3H, α -3L, and α -3H/L, while for α -4, α -4H, α -4L bands were identified. In IEF analysis, a total of 16 bands with independent p I ranging from p I 6.30 to 7.52 were identified for the Glutelin acidic subunit among the cultivars. The maximum and minimum numbers of IEF bands found were 13 and 9, respectively. The α -2L less cultivars were also lacking in p I 6.80 polypeptide in IEF. Result of 2-DE showed that p I 6.80 polypeptide was the main component of α -2L band. Transplanted Aman ecotype was the most diverse with respect to Glutelin variation. Geographical distribution of Glutelin variation in the Transplanted Aman ecotype differed according to the regions. The above results indicate that Bangladesh rice cultivars possess great genetic diversity in Glutelin acidic subunit polypeptides. This study of indigenous rice cultivars from Bangladesh provides useful information regarding their breeding potential.
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the rice mutant esp2 greatly accumulates the Glutelin precursor and deletes the protein disulfide isomerase
Plant Physiology, 2002Co-Authors: Yoko Takemoto, Masahiro Ogawa, Hikaru Satoh, Sean J Coughlan, Thomas W Okita, Toshihiro KumamaruAbstract:Rice (Oryza sativa) accumulates prolamins and Glutelins as storage proteins. The latter storage protein is synthesized on the endoplasmic reticulum (ER) as a 57-kD proGlutelin precursor, which is then processed into acidic and basic subunits in the protein storage vacuole. Three esp2 mutants, CM1787, EM44, and EM747, contain larger amounts of the 57-kD polypeptide and corresponding lower levels of acidic and basic Glutelin subunits than normal. Electron microscopic observation revealed that esp2 contained normal-appearing Glutelin-containing protein bodies (PB-II), but lacked the normal prolamin-containing PB (PB-I). Instead, numerous small ER-derived PBs of uniform size (0.5 μm in diameter) and low electron density were readily observed. Immunoblot analysis of purified subcellular fractions and immunocytochemistry at the electron microscopy level showed that these new PBs contained the 57-kD proGlutelin precursor and prolamin polypeptides. The 57-kD proGlutelin was extracted with 1% (v/v) lactic acid solution only after removal of cysteine-rich prolamin polypeptides, suggesting that these proteins form Glutelin-prolamin aggregates via interchain disulfide bonds within the ER lumen. The endosperm of esp2 mutants contains the lumenal chaperones, binding protein and calnexin, but lacks protein disulfide isomerase (PDI) at the protein and RNA levels. The transcript of PDI was expressed in the seed only during the early stage of seed development in the wild type. These results suggest that PDI plays an essential role in the segregation of proGlutelin and prolamin polypeptides within the ER lumen.
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Improved electrophoretical analyses for rice seed storage Glutelin.
Yi chuan xue bao = Acta genetica Sinica, 2001Co-Authors: Hikaru Satoh, Ogawa MAbstract:Rice seed storage Glutelin could be separated into three acidic (alpha-) and three basic (beta-) subunit components by SDS-PAGE using 15%-25% gradient acrylamide gel. More than 13 acidic and 14 basic bands may be distinguished with an improved iso-electric focus electrophoresis system by adjusting Ampholine ratios. Combine these two sensitive electrophoresis systems resulting in two-dimensional electrophoresis giving high resolution of Glutelin leading the yield of single polypeptide. The improved electrophoresis system is helpful in identifying rice Glutelin variation and for further studies of Glutelin biochemistry.
Kunisuke Tanaka - One of the best experts on this subject based on the ideXlab platform.
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Ultrastructure of low-Glutelin rice endosperm
Plant Biotechnology, 2007Co-Authors: Sachiko Furukawa, Takehiro Masumura, Kunisuke Tanaka, Mitsuaki Itou, Yoshifumi Kiyokawa, Yoshinori WakaiAbstract:Low-Glutelin rice has low content of digestible protein Glutelin, and is expected to use as raw material for sake brewing. To characterize endosperm cell structure of low-Glutelin rice, the storage proteins of endosperm cells in polished rice (degree of milling, 70%; used as raw material for sake brewing) of two low-Glutelin rice varieties, LGC1 and Tashu-kei 1001, and two general rice cultivars (brewer's rice), Hyogokitanishiki and Yamadanishiki, were observed using a transmission electron microscope. Low-Glutelin rice differed from general rice cultivars in the composition of major storage proteins and the transgranular distribution of storage proteins as well as in the distribution of protein bodies in endosperm cells. Furthermore, low-Glutelin rice had a specific endosperm cell structure containing many type I protein bodies even in the outer layer of polished rice grains.
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distribution of storage proteins in low Glutelin rice seed determined using a fluorescent antibody
Journal of Bioscience and Bioengineering, 2003Co-Authors: Sachiko Furukawa, Takehiro Masumura, Kunisuke Tanaka, Yoshifumi Kiyokawa, Tomochika Mizuma, Yoshinori WakaiAbstract:To compare the distribution of storage proteins in low-Glutelin rice seed with that in other cultivars having normal protein compositions, immunofluorescence labeling with specific antibodies was applied to visualize the distribution of storage proteins in endosperm tissues. The endosperm tissues from five cultivars were reacted with anti-prolamin and anti-Glutelin antibodies, and then observed by light microscopy and confocal laser scanning microscopy (CLSM). In low-Glutelin rice, using microscopic analysis, a large proportion of storage proteins was observed in the endosperm tissue of 70% polished rice. To determine the localization of two types of protein bodies in endosperm tissues, images of the distribution of the type I protein body (PB-I) and the type II protein body (PB-II) were obtained by CLSM. The CLSM images showed that, in low-Glutelin rice, prolamin which accumulates in PB-I remains in the center of 70% polished rice grains despite the elimination of 30% of the outer layer of brown rice grains. However, the other cultivars mostly contained Glutelin which accumulates in PB-II and is distributed throughout the endosperm tissues. This shows that low-Glutelin rice differs from the other cultivars not only in the major storage protein composition but also in the distribution of storage proteins in endosperm tissues.
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Molecular Cloning of a Novel Glutelin cDNA from Rice Seeds
Plant Biotechnology, 1998Co-Authors: Norihiro Mitsukawa, Hirofumi Hayashi, Kayo Yamamoto, Kunitomo Kidzu, Ryoichi Konishi, Takehiro Masumura, Kunisuke TanakaAbstract:A novel Glutelin gene was cloned from a cDNA library of maturing rice seeds (Oryza sativa L. cv Nipponbare). The 2.0kbp insert contained an open reading frame encoding a 510 amino acid polypeptide (Mr 57, 116). This novel Glutelin shares 46 to 49% amino acid identity with previously identified rice Glutelins. A phylogenetic analysis of cloned Glutelins indicates that this gene constitutes a new, fifth class of Glutelin gene families. The Asn-Gly processing sequence which is highly conserved in 11S seed storage proteins is replaced by Asn-Val in the sequence of the novel Glutelin. The amino acid composition extending 50 residues both upstream and downstream of the Asn-Val site was less hydrophilic than in other Glutelins. The N-terminal half corresponding to the acidic domains of other Glutelins possesses a higher pI value (7.46) than found in other Glutelins. Expression of the gene was detected in maturing seeds, but not in roots or leaves.