• Journal of Radiation Industry
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• v.2 no.3
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• pp.97-104
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• 2008

Cinnamate-4-hydroxylase (C4H) is a key enzyme of phenylpropanoid pathway, which leads a variety of secondary metabolites to participate in differentiation and protection of plant against environmental stresses. In this study, we isolated a full-length cDNA of the C4H gene from a black raspberry (Rubus occidentalis L.), using a reverse transcriptase-PCR and rapid amplification of the cDNA ends (RACE)-PCR. The full-length cDNA of the RocC4H gene contained a 1,515 bp open reading frame (ORF) encoding a 504 amino acid protein with a calculated molecular weight of about 57.9 kDa and an isoelectric point (pI) value of 9.1. The genomic DNA analysis revealed that RocC4H gene had three exons and two introns. By multiple sequence alignment, RocC4H protein was highly homologous with other plant C4Hs, and the cytochrome P450-featured motifs, such as the heme-binding domain, the T-containing binding pocket motif (AAIETT), the ERR triad, and the tetrapeptide (PPGP) hinge motif, were highly conserved. Southern blot analysis revealed that RocC4H is a single copy gene in R. occidentalis.

• BMB Reports
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• v.41 no.7
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• pp.529-536
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• 2008

Cinnamate-4-hydroxylase (C4H) is a key enzyme in the phenylpropanoid pathway, which is responsible for synthesizing a variety of secondary metabolites that participate in development and adaptation. In this study, we isolated a full-length cDNA of the C4H gene from the Korean black raspberry (Rubus sp.) and found that this gene existed as a single gene. By comparing the deduced amino acid sequence of Rubus sp. C4H with other sequences reported previously we determined that this sequence was highly conserved among widely divergent plant species. In addition, quantitative real time PCR studies indicated that the C4H gene had a differential expression pattern during fruit development, where gene expression was first detected in green fruit and was then remarkably reduced in yellow fruit, followed by an increase in red and black fruit. To investigate the two peaks in expression observed during fruit development and ripening, we measured the flavonoid content. The content of the major flavanol of Korean black raspberry fruits was determined to be highest at the beginning of fruit development, followed by a gradually decrease according to the developmental stages. In contrast, the content of anthocyanins during the progress of ripening was dramatically increased. Our results suggest that the C4H gene in Korean black raspberry plays a role during color development at the late stages of fruit ripening, whereas the expression of C4H gene during the early stages may be related to the accumulation of flavanols.

• Proceedings of the Korean Society of Crop Science Conference
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• 2005.10b
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• pp.120-121
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• 2005

• 한국약용작물학회:학술대회논문집
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• 2011.04a
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• pp.458-459
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• 2011

• 한국약용작물학회:학술대회논문집
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• 2010.10a
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• pp.390-391
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• 2010

• BMB Reports
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• v.40 no.2
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• pp.247-260
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• 2007

Cinnamate 4-hydroxylase (C4H) is a key enzyme of phenylpropanoid pathway, which synthesizes numerous secondary metabolites to participate in development and adaption. Two C4H isoforms, the 2192-bp BnC4H-1 and 2108-bp BnC4H-2, were cloned from oilseed rape (Brassica napus). They both have two introns and a 1518-bp open reading frame encoding a 505-amino-acid polypeptide. BnC4H-1 is 57.73 kDa with an isoelectric point of 9.11, while 57.75 kDa and 9.13 for BnC4H-2. They share only 80.6% identities on nucleotide level but 96.6% identities and 98.4% positives on protein level. Showing highest homologies to Arabidopsis thaliana C4H, they possess a conserved p450 domain and all P450-featured motifs, and are identical to typical C4Hs at substrate-recognition sites and active site residues. They are most probably associated with endoplasmic reticulum by one or both of the N- and C-terminal transmembrane helices. Phosphorylation may be a necessary post-translational modification. Their secondary structures are dominated by alpha helices and random coils. Most helices locate in the central region, while extended strands mainly distribute before and after this region. Southern blot indicated about 9 or more C4H paralogs in B. napus. In hypocotyl, cotyledon, stem, flower, bud, young- and middle-stage seed, they are co-dominantly expressed. In root and old seed, BnC4H-2 is dominant over BnC4H-1, with a reverse trend in leaf and pericarp. Paralogous C4H numbers in Brassicaceae genomes and possible roles of conserved motifs in 5' UTR and the 2nd intron are discussed.

• Journal of Plant Biotechnology
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• v.32 no.3
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• pp.167-173
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• 2005

Three different cDNAS for cinnamate 4-hydroxylase (C4H) which are involved in the second step of the general phenylpropanoid pathway were isolated and designated as pc4h1 (1,755 bp), pc4h2 (1,655 bp), and pc4h3 (1,316 bp), respectively. The nucleotide sequence analysis revealed that both pc4h1 and pc4h2 clones encode polypeptides of 505 amino acids frame but pc4h3 clone was truncated at the 5'-end of coding region. The alignment of the deduced amino acid sequences showed that PC4H1 and PC4H2 are highly homologous (95.8% identical) with each other and contain three conserved domains which are typical in cytochrome P450 monooxygenase: proline-rich region, threonine-containing binding pocket for the oxygen molecule, and heme binding region. In addition, result of the phylogenic tree analysis revealed that both pepper C4Hs belong to Class 1. pc4h2 transcription was strongly induced in wounded fruit (400%) and root (200%) relative to its very low basal level but not in leaf or stem tissue. In case of pc4h1, the basal level of transcription was higher than pc4h2 but induction by wounding was lower in fruit and root while leaf and stem tissues did not respond to wounding. The basal level of pc4h3 transcripts was not, if any, detectable and response to wounding was not observed.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2005.04a
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• pp.59-59
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• 2005

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2005.04a
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• pp.107-107
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• 2005

• Journal of Plant Biology
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• v.37 no.2
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• pp.223-229
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• 1994

장미(Rosa sp. cv. Paul's Scarlet)와 밀(Triticum aestivum L.)의 조직 배양체에서 polychlorinated biphenyl(PCB)의 대사를 조사하였다. 조사된 PCB는 2개에서 6개까지의 치환 염소를 보유한 19종류였다. PCB를 투여하여 96시간 동안 배양시킨 결과 장미에서는 9종, 밀에서는 5종류가 30% 이상의 대사율을 나타내었다. 대사율이 높은 PCB들은 모두 2번 위치에 치환 염소를 갖는다는 공통점이 있으며, 대사율이 낮은 PCB일수록 평면적 구조를 갖는 것으로 나타났다. 한편 밀의 배양세포는 para- 위치에 치환 염소를 갖는 PCB에 대한 대사 활성이 전혀 없었다. 두 종의 배양 세포 모두에서 phenobarbital을 처리한 경우 non-p-chlorinated biphenyl의 대사율만이 증가하였으며, parachlorinated biphenyl의 대사 활성은 phenobarbital 처리에 영향을 받지 않았다. 또한 phenobarbital의 처리에 의하여 cinnamate-4-hydroxylase의 활성이 140% 이상 증가하였다. 이상의 결과는 식물체에서의 PCB 대사가 그 구조에 의해 결정될 수 있으며, 특히 para- 위치의 치환 염소를 보유한 종류와 그렇지 않은 종류는 별개의 cytochrome p450의 동위 효소에 의하여 대사될 수 있음을 보여주고 있다.