N-Acetyl-d-neuraminic acid (Neu5Ac) is
a potential baby nutrient and the key precursor of antiflu medicine
Zanamivir. The Neu5Ac chemoenzymatic synthesis consists of N-acetyl-d-glucosamine epimerase (AGE)-catalyzed
epimerization of N-acetyl-d-glucosamine
(GlcNAc) to N-acetyl-d-mannosamine (ManNAc)
and aldolase-catalyzed condensation between ManNAc and pyruvate. Herein,
we cloned and characterized BT0453, a novel AGE, from a human gut
symbiont Bacteroides thetaiotaomicron. BT0453 shows
the highest soluble fraction among the AGEs tested. With GlcNAc and
sodium pyruvate as substrates, Neu5Ac production by coupling whole
cells expressing BT0453 and Escherichia coli N-acetyl-d-neuraminic acid aldolase was explored. After 36 h, a 53.6%
molar yield, 3.6 g L–1 h–1 productivity
and 42.9 mM titer of Neu5Ac were obtained. Furthermore, for the first
time, the T7-BT0453-T7-nanA polycistronic
unit was integrated into the E. coli genome, generating
a chromosome-based biotransformation system. BT0453 protein engineering
and metabolic engineering studies hold potential for the industrial
production of Neu5Ac.
Background
The redundancy of genomic resources, including transcript and molecular markers, and their uncertain position in the genome have dramatically hindered the study of traits in ramie, an important natural fiber crop.
Results
We obtained a high-quality transcriptome consisting of 30,591 non-redundant transcripts using single-molecule long-read sequencing and proposed it as a universal ramie transcriptome. Additionally, 55,882 single nucleotide polymorphisms (SNPs) were identified and a high-density genetic map was developed. Based on this genetic map, 181.7 Mb ramie genome sequences were assembled into 14 chromosomes. For the convenient use of these resources, 29,286 (~ 95.7%) of the transcripts and all 55,882 SNPs, along with 1827 previously reported sequence repeat markers (SSRs), were mapped into the ramie genome, and 22,343 (~ 73.0%) transcripts, 50,154 (~ 89.7%) SNPs, and 1466 (~ 80.3%) SSRs were assigned to a specific location in the corresponding chromosome.
Conclusion
This is the first study to characterize the ramie transcriptome by long-read sequencing, and the substantial number of transcripts of significant length obtained will accelerate our understanding of ramie growth and development. This integration of genome sequences, expressed transcripts, and genetic markers will provide an extremely useful resource for genetic, molecular, and breeding studies of ramie.
Electronic supplementary material
The online version of this article (10.1186/s12864-019-5878-8) contains supplementary material, which is available to authorized users.
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