Week 2 HW

Part 1: Define your expression target

What molecule are you expressing? I want to express the enzyme urate oxidase to aid in the breakdown of uric acid, the end metabolite of purine metabolism.

What is the sequence? The protein sequence for uricase will be taken from Vibrio vulnificus and can be found from NCBI accession number: TDL85923.1

MTIQNENRTMFYGKGDVFVYRTFVQPLTGLRKIPESSFTERDNTIFGFNCQISLKGEAFFSSFTEGDNSL VVATDSMKNIIHRQAANYQGNTAEGFLKFICEVFLDKYSHIDAVELTATEVPFDHVLVPKGEGHENSDVV YRCSRNERATTTIEVKRTPTGSKIVKHSSGIVDVHLIKVKGSSFYGFIQDEYTTLPEAQDRPLFIYLDFD WEYSRWEDGTGANPEKYVAAEQICDIANTVFHELNNRSIQQLIYHIGIRILERFPQLANVQFKTNNRTWE TVVETIPNSEGSVYQEPRPPFGFQGFVVTQEDVKQKKAADSTVGASR

What host or chassis? I want to use a probiotic organism to act as a transient gut microbe to help relieve hyperuricemia. For this, I would use Saccharomyces boulardii that already has applications as a biotherapeutic probiotic and is typically cleared from the gut within 2-5 days post-supplementation.

Where does the product need to go? The product will be expressed intracellularly in the cytoplasm of Saccharomyces boulardii. In order for it to come in contact with uric acid, I will have to additionally express a urate transporter (Such as UapA)

References:

  1. Wenzhuo Wang, Lei Pan, Huansha He, Huiyuan Xue, He Huang, Audrey Mihewi Samosir, Xian Fu, Yue Shen; Systematic Engineering for Efficient Uric Acid-Degrading Activity in Probiotic Yeast Saccharomyces boulardii. ACS Synth. Biol. 20 June 2025; 14 (6) 2030 2043. https://doi.org/10.1021/acssynbio.4c00831
  2. National Center for Biotechnology Information. (n.d.). Urate oxidase [Vibrio vulnificus] (Protein accession No. TDL85923.1). NCBI Protein. NCBI Protein record

Part 6. What dont you know yet?

My biggest unknown at the moment is whether the host system and transporter protein will be functional at a 37°C body temperature as opposed to their preferred 30°C Additionally, since this probiotic is mainly meant to reduce the side effects of high nucleic acid single cell proteins, I wonder if administering both at the same time would be optimal for uric acid removal.