De Novo Synthesis of Perdeuterated Phosphoinositide by Installing a Non-native Phospholipid Biopathway in E. coli

TY - JOUR

T1 - De Novo Synthesis of Perdeuterated Phosphoinositide by Installing a Non-native Phospholipid Biopathway in E. coli

AU - Merrild, Aske Høj

AU - Johnsen, Niels Krabbe

AU - Zhang, Mingliang

AU - Bogojevic, Oliver

AU - Ouyang, Yi

AU - Guo, Zheng

PY - 2024/10/18

Y1 - 2024/10/18

N2 - Phosphatidylinositol (PI) and its phosphorylated derivatives are of paramount importance in cellular functions and diseases. Understanding their diverse roles is, however, challenged by difficulties in synthesis and labeling techniques. In this proof-of-concept study, we demonstrate that PI can be straightforwardly de novo-synthesized and deuterium (2H)-labeled in Escherichia coli by genomic insertion of PI synthase from Trypanosoma brucei under constitutive synthetic promoter proD. Insertion into loci atpi-gidB and ybb revealed PI accumulation of 41% and 34% (mol/mol), respectively, when cultivated with glycerol as the sole carbon source. Growth of the atpi-gidB-PIS strain in deuterium-labeled (2H) substrates D2O, D8-glycerol, and D6-myo-inositol achieved PI deuteration of 90%, PE deuteration of 95%, and total fatty acids|fatty acid (FA) deuteration of 97%. This study offers an alternative convenient route to chemical and enzymatic labeling synthesis of PI; more excitingly, this work also, in principle, opens a door for tailoring the FA profile of deuterated PI/PE for task-specific application by repurposing FA biosynthesis pathways. (Figure Presented).

AB - Phosphatidylinositol (PI) and its phosphorylated derivatives are of paramount importance in cellular functions and diseases. Understanding their diverse roles is, however, challenged by difficulties in synthesis and labeling techniques. In this proof-of-concept study, we demonstrate that PI can be straightforwardly de novo-synthesized and deuterium (2H)-labeled in Escherichia coli by genomic insertion of PI synthase from Trypanosoma brucei under constitutive synthetic promoter proD. Insertion into loci atpi-gidB and ybb revealed PI accumulation of 41% and 34% (mol/mol), respectively, when cultivated with glycerol as the sole carbon source. Growth of the atpi-gidB-PIS strain in deuterium-labeled (2H) substrates D2O, D8-glycerol, and D6-myo-inositol achieved PI deuteration of 90%, PE deuteration of 95%, and total fatty acids|fatty acid (FA) deuteration of 97%. This study offers an alternative convenient route to chemical and enzymatic labeling synthesis of PI; more excitingly, this work also, in principle, opens a door for tailoring the FA profile of deuterated PI/PE for task-specific application by repurposing FA biosynthesis pathways. (Figure Presented).

KW - labeled lipids

KW - perdeuterated lipids

KW - phosphatidylinositol synthase

KW - phosphoinositol

KW - phospholipids

UR - http://www.scopus.com/inward/record.url?scp=85204438369&partnerID=8YFLogxK

U2 - 10.1021/acssynbio.4c00413

DO - 10.1021/acssynbio.4c00413

M3 - Journal article

C2 - 39292964

SN - 2161-5063

VL - 13

SP - 3344

EP - 3353

JO - ACS Synthetic Biology

JF - ACS Synthetic Biology

IS - 10

M1 - 39292964

ER -