plant 2차 대사산물 - 치료적 식물 탐구!!

[문형철]

Since ages plants with therapeutic effects are used for curing several human ailments. With the development in modern biotechnological tools, these therapeutic compounds could be overexpressed in the plants. This review aims to compile the valuable information from the examples in the literature, regarding the biotechnological tools those could be useful for the overproduction of health-promoting biochemicals in the medicinal plants. Gene level modification is all about creating improved variety of plants that are highly resistant to pests and pesticides or contain higher levels of nutrients than conventional plants. Plant secondary metabolites constitute an exciting and vital aspect of research, due to their chemical diversity, varied biological functions and pharmacological activities. Gene modification, through Agrobacterium mediated transformation, RNA interference or CRISPER/Cas9, has opened avenues of improvement in medicinal plants and provide an alternative production system of pharmaceutically important secondary metabolites for their commercial exploitation.

오랫동안 치료 효과가 있는 식물은 여러 가지 인간의 질병을 치료하는 데 사용되어 왔습니다. 현대 생명공학 도구의 발달로 이러한 치료 화합물을 식물에 과다 발현시킬 수 있게 되었습니다. 이 리뷰는 약용 식물에서 건강에 좋은 생화합물을 과다 생산하는 데 유용할 수 있는 생명공학 도구에 관한 문헌의 사례에서 얻은 귀중한 정보를 수집하는 것을 목표로 합니다. 

유전자 수준 변형은 

해충과 살충제에 대한 저항성이 높거나 

기존 식물보다 더 많은 양의 영양소를 함유하는 

개선된 품종의 식물을 만드는 것입니다. 

식물 이차 대사 산물은 

화학적 다양성, 다양한 생물학적 기능, 약리학적 활동으로 인해 

흥미롭고 중요한 연구 대상입니다.

 유전자 변형은 A

grobacterium 매개 형질전환, 

RNA 간섭 또는 CRISPER/Cas9를 통해 

약용 식물의 개선 가능성을 열어주었고, 

상업적 이용을 위한 의약적으로 중요한 이차 대사 산물의 대체 생산 시스템을 제공합니다.

INTRODUCTION

Plants synthesize innumerable small phytomolecules, known metabolites via integrated complex interconnected enzyme-mediated biosynthetic pathway.1 Plants primarily synthesize two types of metabolites: The primary metabolites, which are vital for the survival of the organisms and any change in their synthesis, can lead to harmful manifestation.2 However, secondary metabolites confer a multitude of adaptive and evolutionary advantages to the plants by performing specialized functions and playing a role in providing quality of life to the producer [Figure 1]. Often, secondary metabolites are associated with defense related, antifeedent, insect attractant and repellent functions.3,4 Plants are the richest source for the secondary metabolites, which have medicinal and aromatic properties. According to some estimates, approximately 100,000 such secondary metabolites are known to occur in about 50,000 plant species and more than 4,000 unique metabolic compounds are being found each year from several plant species.5-8 For thousands of years, we have been dependent on natural plant products for human healthcare in the form of drugs, antioxidants, flavors, fragrances, dyes, insecticides and pheromones.9,10 The global plant-derived product market is expected to gain momentum over the coming years.11 Firstly, due to the low cost of herbal medicines compared to allopathy and secondly, due to fewer side effects and no overdose toxicity, that are so common in the case of synthetic drugs. There is an increasing inclination of consumers towards traditional medicines.

Consequently, global herbal medicine (Ayurveda, Unani and Traditional Chinese Medicine) market size which was valued at USD 71.19 billion in 2016, is expected to reach $ 111 billion by the end of 2023. This growth in market, is due to increasing demand and can only be met through intensive research investments and funding, which will support constant supply of these herbal products in near future.12 Despite such a substantial increase in demand for nutraceuticals, the associated growth in medicinal crop cultivation has marginally increased. The reasons may be identified as (1) low concentration of targeted phytomolecules in specific plant tissues, (2) technological neglect in growing medicinal crops, (3) conversion of habitats of medicinal plants to food or cash crop cultivation, (4) conventional breeding techniques do not provide desirable level of improvement due to sterility, long generation time and complex biosynthetic pathways involved.13,14 In this review, we have compiled the useful information after extensive literature survey concerning the biotechnological tools and those could be useful for the overproduction of health-promoting biochemicals in the medicinal plants

식물은 

통합된 복잡한 상호 연결된 효소 매개 생합성 경로를 통해 

수많은 작은 식물 분자, 

즉 대사 산물을 합성합니다.1 

식물은 주로 

두 가지 유형의 대사 산물을 합성합니다. 

1차 대사 산물은 

유기체의 생존과 합성 변화에 필수적이며, 

유해한 증상을 유발할 수 있습니다.2 

그러나 

2차 대사 산물은 

특수한 기능을 수행하고 

생산자에게 삶의 질을 제공하는 역할을 함으로써 

식물에 다양한 적응 및 진화적 이점을 제공합니다[그림 1]. 

종종, 

2차 대사 산물은 

방어 관련, 항식충, 곤충 유인 및 방충 기능과 관련이 있습니다.3,4 

식물은 

약용 및 향기로운 특성을 지닌 

2차 대사 산물의 가장 풍부한 공급원입니다. 

일부 추정에 따르면, 

약 50,000종의 식물에서 

약 100,000개의 2차 대사 산물이 생성되는 것으로 알려져 있으며,

 매년 여러 식물 종에서 4,000개 이상의 독특한 대사 화합물이 발견되고 있습니다.5-8 

수천 년 동안 우리는 

약, 항산화제, 향료, 향수, 염료, 살충제, 페로몬 등의 형태로 

자연 식물 제품을 인간의 건강 관리에 의존해 왔습니다.9,1 0 

전 세계 식물 유래 제품 시장은 

향후 몇 년 동안 성장세를 보일 것으로 예상됩니다.11 

첫째, 동종 요법에 비해 한약의 비용이 저렴하고, 

둘째, 합성 약물의 경우 흔히 나타나는 부작용이 적고 과다 복용으로 인한 독성이 없다는 점 때문입니다. 

소비자의 전통 의약품 선호도가 증가하고 있습니다.

따라서 

2016년 711억 9천만 달러로 평가된 글로벌 한방 의약품(아유르베다, 우나니, 전통 중국 의학) 시장 규모는 

2023년 말까지 1,110억 달러에 이를 것으로 예상됩니다. 

이러한 시장의 성장은 

수요 증가에 기인하며, 

집중적인 연구 투자와 자금 지원을 통해서만 충족될 수 있으며, 

이는 가까운 미래에 이러한 허브 제품의 지속적인 공급을 지원할 것입니다.12 

이러한 기능성 식품에 대한 수요가 크게 증가했음에도 불구하고, 

약용 작물 재배의 관련 성장은 미미하게 증가했습니다. 

그 이유는 

(1) 특정 식물 조직에서 표적 식물 분자의 농도가 낮음, 

(2) 약용 작물 재배에 대한 기술적 방치, 

(3) 약용 식물의 서식지를 식량 또는 현금 작물 재배로 전환, 

(4) 불임, 

긴 세대 시간 및 복잡한 생합성 경로로 인해 기존 육종 기술이 바람직한 수준의 개선을 제공하지 못함으로 추정할 수 있습니다.13,14 

이 리뷰에서는 광범위한 문헌 조사를 통해 

생명 공학 도구에 관한 유용한 정보를 수집했습니다. 

그리고 

그것들은 

약용 식물에서 건강에 좋은 생화합물을 과다 생산하는 데 

유용할 수 있습니다.

GENE LEVEL MODIFICATION THROUGH METABOLIC ENGINEERING

To increase the production of (a group of) compound(s), directed and user manipulation of metabolic pathways at gene level in a living system can be performed by manipulating transporters along with enzymatic and regulatory functions of the cell15,16 and eggplant fruits are of different shape and sizes that render them as an ideal system for metabolic engineering. Here, we have developed an agroinfiltration protocol for the transient expression‎ of a gene in the eggplant fruit using GUS bearing; pCAMBIA1304 vector. Thereafter, to prove the effectiveness of the developed protocol, we have used the eggplant hydroxycinnamoyl CoA-quinate transferase (SmHQT Metabolic engineering is now used to improve medicinal plants to meet the requirements of increased human health.17 Metabolic engineering may be approached through, overexpression‎ or upregulation of gene(s) specific to rate-limiting steps in the pathway and by blocking competitive pathways to reduce degradation of the product of interest.18

There have been efforts to alter the expression‎ of some regulatory genes that control multiple genes in the biosynthetic pathway, like down regulation of zeaxanthin epoxidase gene by antisense and co-suppression inhibition, which resulted in an accumulation of zeaxanthin.19 By means of metabolic engineering, selective novel metabolites can be produced, the target of secondary metabolite can be overproduced and proportion of toxic and unwanted chemicals can be reduced. The strategy to amplify the metabolic flux of a pathway can be achieved by increasing the composition of the enzyme involved in the rate-limiting step of the pathway.20 With progress in tissue culture techniques, combined with development in genetic engineering and bioinformatics, have opened a panorama for large-scale and improved production of secondary metabolites with pharmaceutical and nutritional value.21

(한 그룹의) 화합물의 생산을 늘리기 위해, 

살아있는 시스템의 유전자 수준에서 대사 경로의 

직접적, 사용자 조작이 세포의 효소 및 조절 기능과 함께 

수송체를 조작함으로써 수행될 수 있습니다15,16 

가지 과일은 

모양과 크기가 다양하여 대사 공학에 이상적인 시스템입니다. 

여기서는 GUS를 보유한 pCAMBIA1304 벡터를 사용하여 

가지 과일에서 유전자를 일시적으로 발현하는 농작물 침투 프로토콜을 개발했습니다. 

그 후, 개발된 프로토콜의 효과를 증명하기 위해 

가지 하이드록시신나모일 CoA-퀴네이트 전이효소(SmHQT)를 사용했습니다. 

대사 공학은 이제 인간의 건강 증진이라는 요구 사항을 충족시키기 위해 약용 식물을 개선하는 데 사용되고 있습니다.17 대사 공학은 경로의 속도 제한 단계에 특정한 유전자(들)의 과발현 또는 상향 조절을 통해 접근할 수 있으며, 경쟁 경로를 차단하여 관심 대상 제품의 분해를 줄일 수 있습니다.18

생합성 경로에서 여러 유전자를 조절하는 일부 조절 유전자의 발현을 변화시키는 노력이 있었습니다. 

예를 들어, 

제아잔틴 에폭시다제 유전자의 다운레귤레이션과 

코억제 억제를 통해 

제아잔틴이 축적되는 결과를 가져왔습니다.19 

대사 공학을 통해 선택적 신진대사 산물을 생산할 수 있고, 

이차 대사 산물의 표적을 과잉 생산할 수 있으며, 

독성 및 원치 않는 화학 물질의 비율을 줄일 수 있습니다. 

경로의 대사 흐름을 증폭시키는 전략은 

경로의 속도 제한 단계에 관여하는 효소의 구성을 증가시킴으로써 달성될 수 있습니다.20 

조직 배양 기술의 발전과 유전공학 및 생물정보학의 발전이 결합되어, 

의약 및 영양적 가치가 있는 

2차 대사 산물의 대규모 생산과 개선된 생산을 위한 파노라마가 열렸습니다.21

STRATEGIES TO INDUCE GENE LEVEL MODIFICATION

Various genetic manipulation techniques and technologies have been applied in plants, such as overexpression‎ of transgenes, expressions of multiple transgenes, gene silencing and overproduction of bioactive plant natural products.22,23 To mention a few, three most commonly used methods are discussed below: AGROBACTERIUM MEDIATED TRANSFORMATION For transferring useful genes into crop plants, Agrobacterium-mediated gene transfer is the most frequently used method of transformation, either in differentiated plant cells or into undifferentiated cells of callus [Figure 2]. This method uses the natural ability of Agrobacterium to infect and transfer genes to the plant cells. The success of this method relies on many factors, like type of target cells or tissues and its competence for regeneration and transformation, efficiency of DNA delivery, stringency of system used for selection of transformed cells and the ability to recover fertile transgenic plants.24]

Agrobacterium tumefaciens mediated transformation systems in medicinal crops is pertinent to highly efficient and rapid transgenic plant production for desirable traits for cultivation. Increased yield, biotic and abiotic stress tolerance, hyper-production of one or more desirable secondary metabolite or other phytochemicals, suppression of synthesis of one or more undesirable phytochemical, suppression of one or more native genes, metabolic engineering of endogenous pathways, incorporation of genetic traits (genes) of new or non-native metabolic pathways in the plant etc.,25,26 A. rhizogenes-based transformation is another potential system for manipulation in biosynthesis of secondary metabolites, mainly in roots of medicinal plants. Ri transformed hairy root cultures shows rapid growth, reduced apical dominance, high branching and increased large scale production of targeted secondary metabolites.27

유전자 수준 변형을 유도하는 전략

식물에는 트랜스유전자의 과발현, 다중 트랜스유전자의 발현, 유전자 침묵화, 생리활성 식물 천연물의 과잉 생산 등 다양한 유전자 조작 기법과 기술이 적용되어 왔습니다.22,23 몇 가지를 언급하자면, 가장 일반적으로 사용되는 세 가지 방법이 아래에 설명되어 있습니다. AGROBACTERIUM MEDIATED TRANSFORMATION 유용한 유전자를 작물 식물에 옮기기 위해, Agrobacterium-mediated gene transfer는 분화 식물 세포 또는 미분화 세포인 캘러스에 유전자를 옮기는 가장 자주 사용되는 변형 방법입니다 [그림 2]. 이 방법은 Agrobacterium이 식물 세포를 감염시키고 유전자를 옮기는 자연적인 능력을 이용합니다. 이 방법의 성공은 표적 세포나 조직의 유형, 재생과 변형 능력, DNA 전달 효율, 변형된 세포 선택에 사용되는 시스템의 엄격함, 그리고 비옥한 형질전환 식물을 복구하는 능력 등 여러 가지 요소에 달려 있습니다.24]

약용 작물에서 Agrobacterium tumefaciens 매개 형질전환 시스템은 재배에 적합한 형질을 가진 형질전환 식물을 매우 효율적이고 빠르게 생산하는 것과 관련이 있습니다. 수확량 증가, 생물적 및 비생물적 스트레스 내성, 하나 이상의 바람직한 2차 대사 산물 또는 기타 식물 화학 물질의 과다 생산, 하나 이상의 바람직하지 않은 식물 화학 물질의 합성 억제, 하나 이상의 고유 유전자 억제, 내인성 경로의 대사 공학, 식물 내 새로운 또는 비 고유 대사 경로의 유전적 특성(유전자) 통합 등,25,26 A. rhizogenes 기반 형질전환은 주로 뿌리에서 2차 대사 산물의 생합성을 조작할 수 있는 또 다른 잠재적 시스템입니다. 약용 식물의. Ri 변형 털이 많은 뿌리 배양은 빠른 성장, 감소된 정점 우성, 높은 가지 형성, 표적 2차 대사 산물의 대규모 생산 증가를 보여줍니다.27