Recent Advances in Metabolic Engineering, Protein Engineering, and Transcriptome-Guided Insights Toward Synthetic Production of Taxol

Abstract

The diterpenoid paclitaxel (Taxol®) is a blockbuster anticancer agent that was originally isolated from the Pacific yew (Taxus brevifolia) five decades ago. Despite the wealth of information gained over the years on Taxol research, there still remains supply issues to meet increasing clinical demand. Although alternative Taxol production methods have been developed, they still face several drawbacks that cause supply shortages and high production costs. It is highly desired to develop biotechnological production platforms for Taxol, however, there are still gaps in our understanding of the biosynthetic pathway, catalytic enzymes, regulatory and control mechanisms that hamper production of this critical drug by synthetic biology approaches. Over the past 5 years, significant advances were made in metabolic engineering and optimization of the Taxol pathway in different hosts, leading to accumulation of taxane intermediates. Computational and experimental approaches were leveraged to gain mechanistic insights into the catalytic cycle of pathway enzymes and guide rational protein engineering efforts to improve catalytic fitness and substrate/product specificity, especially of the cytochrome P450s (CYP450s). Notable breakthroughs were also realized in engineering the pathway in plant hosts that are more promising in addressing the challenging CYP450 chemistry. Here, we review these recent advances and in addition, we summarize recent transcriptomic data sets of Taxus species and elicited culture cells, and give a bird’s-eye view of the information that can be gleaned from these publicly available resources. Recent mining of transcriptome data sets led to discovery of two putative pathway enzymes, provided many lead candidates for the missing steps and provided new insights on the regulatory mechanisms governing Taxol biosynthesis. All these inferences are relevant to future biotechnological production of Taxol.

The Epigenetic Regulation in Plant Specialized Metabolism: DNA Methylation Limits Paclitaxel in vitro Biotechnological Production

“…Nevertheless, some epigenetic variations have been proved to be meiotically heritable in plants without altering the DNA sequence (Kaeppler and Phillips, 1993), later defining the so-called epialleles. This distinctive mechanism supports the existence of an epigenetic memory in plant genomes, although the specific mechanisms are unexplored yet (Mutanda et al, 2021).…”

“…This natural antitumoral agent is a diterpenoid specialized metabolite that was first isolated in the late '60s. Paclitaxel blocks the mitosis progression and the activation of the mitosis checkpoint, leading to apoptosis or to the reversion of cancer cells to G0 phase (Mutanda et al, 2021). Its biosynthetic pathway comprises 19 enzyme-catalyzed steps, 15 of which are known, starting from the precursor geranylgeranyl diphosphate (GGPP) and leading to paclitaxel itself (Croteau et al, 2006).…”

PyMiner: A method for metabolic pathway design based on the uniform similarity of substrate-product pairs and conditional search

“…Taxadiene is a crucial intermediate metabolite in the biosynthetic pathway of paclitaxel. As an important anti-cancer drug, paclitaxel is widely applied in the treatment of ovarian cancer, breast cancer and non-small cell lung cancer [ 43 ]. As shown in Fig 7 , the biosynthesis pathway of taxadiene introduced to S .…”

Role of female-predominant MYB39-bHLH13 complex in sexually dimorphic accumulation of taxol in Taxus media

“…The taxol biosynthesis pathway consists of many functional genes and regulators [ 29 ]. To date, most of the functional genes involved in the taxol biosynthesis pathway have been identified [ 30 ].…”