New preprint on yeast bioengineering to produce modular plant hemicellulose

Despite rising interest in the engineering of polysaccharide-based biomaterials, relatively little is known about how plant hemicelluloses can be built and tailored in non-plant cell factories. Pichia pastoris is a promising host to study cellulose synthase-like A (CSLA; heteromannan synthases) and CSLC (xyloglucan synthase) activities but the functions of specific protein motifs and the impact of their products remain unclear. Here, we used a Golden Gate-based system to assemble chimeric CSL enzymes and optimize yeast cultivation to rapidly produce hemicellulose in mg scale.

Part of Figure 1: Schematic of two plant CSLA enzymes, whose protein domains were exchanged in this study to generate a series of chimeric enzymes.

Interestingly, prolonged expression of a konjac AkCSLA3 glucomannan synthase was toxic to yeast cells, but this impairment was restored by swapping its C-terminal region with that of an Arabidopsis AtCSLA2 mannan synthase. Additional chimeras exceeded the yields of the native enzymes. Overall, cell size was increased or decreased depending on the CSLA sequences used.

Part of Figure 2. Mannose (Man) content, representing insoluble heteromannans, is significantly increased in three of the eight chimeric enzymes tested compared to the parental controls.

Part of Figure 5: The growth rate of the AkCSLA3 yeast strain is significantly lower than the rest. Prolonged glucomannan synthase expression also reduces cell integrity, evidenced by the uptake of the trypan blue strain (panel b), and size.

Yeast containing plant-like polysaccharides could be viewed as tunable biological capsules for further metabolic engineering studies. Sensitive macromolecules such as therapeutic proteins can be protected by encapsulation in non-toxic plant polysaccharides and Pichia cells are attractive hosts for recombinant protein production. Therefore, our strategy provides basic insight into matrix polysaccharide biosynthesis and could also be used to encapsulate valuable cargo.

Publication:

M Robert, J Waldhauer, F Stritt, B Yang, M Pauly, C Voiniciuc (2021) Rapid, modular biosynthesis of plant hemicellulose and its impact on yeast cells bioRxiv, https://doi.org/10.1101/2021.04.20.44061

Two new MSc students join the lab

We are happy to welcome Sooyun Lee and Tilman Jacob to the Designer Glycans lab. Sooyun first joined us for an internship in February to evaluate fluorescent reporters in Pichia pastoris. She will now start her MSc research thesis on secreting proteins from yeast to modify extracellular glycans.

Tilman is a first-year MSc student at the Martin Luther University, who will initially work as a research assistant. Having previously worked with G-protein-coupled receptors in human cell lines, he is not afraid to investigate some our of membrane-bound glycosyltransferases.

View their profiles on The Team page.

Register now for the Leibniz Symposium on Plant Cell Walls

Debora Gasperini, Katharina Bürstenbinder and Catalin Voiniciuc are co-organizing the Leibniz Plant Biochemistry Symposium 2021. This annual conference will be focused on Plant Cell Walls this year, and will be held as an online event for the first time.

We have an exciting speaker line-up and there will be free registration for participants from around the world. Space is limited, so don’t miss out.

Register now: https://webcast.meetyoo.de/reg/dZnJ5vMwozrM

Symposium website: Link

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February publication update

We are happy to have contributed to a new publication that appeared in Science Advances on the February 10. This study unveils how jasmonate signaling is initiated in the korrigan1 (kor1) cellulose-deficient mutant, and multiple ways in which it can be complemented. The work was lead by Stefan Mielke in Debora Gasperini’s research group at IPB Halle.

Article: https://doi.org/10.1126/sciadv.abf0356

Catalin Voiniciuc performed the cell wall analyses, and would like to acknowledge the valuable technical assistance of Christine Wagner and Bo Yang. The Science Advances study was originally available as a pre-print:

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February also marks the release of a correction to the Voiniciuc et al. (2018) Plant Physiology study of pectin biosynthetic enzymes. Unfortunately, there was an accidental mix-up of GAUT11 and another GAUT protein expressed at the same time. Catalin would like to thank the co-authors for reporting the problem affecting Figure 5 and Supplemental Figure 4A, and repeating the experiments. The corrigendum has addressed this issue and shows that the original results and conclusions that GAUT11 is a galacturonosyltransferase (GalAT) that elongates homogalacturonan (HG) are unaffected.

Original article: https://doi.org/10.1104/pp.18.00584

Corrigendum: https://doi.org/10.1093/plphys/kiaa037

New publication in New Phytologist

Our revised manuscript on the roles of branched xylan and heteromannan (previously available as a preprint) in controlling seed mucilage properties and early growth in salt stress has now been peer-reviewed.

Read the open-access article here: https://doi.org/10.1111/nph.17056