Part:BBa_K302010

Svp 4'-Phosphopanthetheinyl-transferase

svp encodes for a 4'-Phosphopanthetheinyl-transferase (PPTase) which transfers a 4'-PPT residue from CoA to a conserved serine residue in the T-Domain of NRPS modules, thus activating these enzymes.

Figure 1: General mechanism of activation the activation of NRP Synthetases from apo- to holo-form.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 173
  Illegal NgoMIV site found at 413
  Illegal NgoMIV site found at 663
  Illegal AgeI site found at 573
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 366
  

References

• Takahashi, H., Kumagai, T., Kitani, K., Mori, M., Matoba, Y., & Sugiyama, M. (2007). Cloning and Characterization of a Streptomyces Single Module Type Non-ribosomal Peptide Synthetase Catalyzing a Blue Pigment Synthesis *, 282(12), 9073–9081. doi:10.1074/jbc.M611319200

Information contributed by City of London UK (2021)

Part information is collated here to help future users of the BioBrick registry.

Metadata:

• Group: City of London UK 2021
• Author: Janusan Jeyananthan
• Summary: Added information collated from existing scientific studies

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Kinetics:

kcat is 86 min(-1) with S.glaucescens apo-ACP TcmM as the substrate and 11 min(-1) with S.mobaraensis apo-PCP BlmI as the substrate.

For Streptomyces mobaraensis apo-PCP BlmI1: KM=3.9 µM For Streptomyces glaucescens apo-ACP TcmM: KM=3.1 µM

Information contributed by NFLS_Nanjing (2022)

Part information is collated here to help future users of the BioBrick registry.

Metadata:

• Group: NFLS_Nanjing (2022)
• Author: Yuming Xue
• Summary: Added information collated from existing scientific studies

Biology and usage:

1. The gene bpsA is expressed in C. glutamicum and carries strong fluxes toward L-glutamate, a precursor of indigoidine. [1]

2. The gene bpsA catalyzes indigoidine formation from two molecules of glutamine in an ATP-dependent manner.[2]

Experimental approach:

1. Integrated sfp into the yeast chromosomal δ-sequences.

2. Codon-optimize Te bpsA gene for expression in S. cerevisiae and genomically integrated into locus ARS1014a under control of the TDH3 promoter and ADH1 terminator using a previously reported, cloning free Cas9 toolkit.

3. Use the conventional lithium acetate method to perform transformations.

4. Use 200 ng pCut_1014a and 500 ng of linear Donor DNA with 500 bp homology to the integration locus ARS1014a.[3]

References

[1] High-Level Production of the Natural Blue Pigment Indigoidine from Metabolically Engineered Corynebacterium glutamicum for Sustainable Fabric Dyes [2] Genome-scale metabolic rewiring improves titersrates and yields of the non-native product indigoidine at scale

[3] Production efciency of the bacterial non-ribosomal peptide indigoidine relies on the respiratory metabolic state in S. cerevisiae