Device

Part:BBa_K4593021

Designed by: Jianfei Song   Group: iGEM23_BNDS-China   (2023-10-08)
Revision as of 00:12, 9 October 2023 by Jianfei Song (Talk | contribs) (→‎QS-detection circuit)


S. aureus in vivo elimination apparatus for B. subtilis

Overview

This part contains 3 genetic circuits: an S. aureus quorum sensing (QS) detection system (BBa_I746101), an Endolysin combinative expression system (BBa_K4593018), and a self-lysing release system (BBa_I746104 and BBa_K4593001).

As the modified final intended product of our project's S. aureus elimination module, this device could constitutively express S. aureus targeting endolysins (a set of enzymes that specifically lyses S. aureus) and detect the presence of S. aureus QS signal. If the QS signal molecule is detected, the self-lysing enzyme composite (Spn1s_LysRZ) will be activated, and the engineered cell will lyse itself to release the endolysin that eliminates pathogenic S. aureus.

Endolysin expression circuit

This circuit could constitutively express 3 types of S. aureus targeting endolysin-LysDZ25, ClyC, and Lys GH15-that have maximum efficiency conditions complementary with each other. This ensures that at least one type of endolysin will be functional under various conditions in the human digestive tract. The lytic efficiency of the endolysins with respect to concentration is being characterized separately in our project. For detailed information, see the following basic parts:

LysDZ25: BBa_K4593000

ClyC: BBa_K4593002

LysGH25: BBa_K4593003

QS-detection circuit

This circuit could constitutively express AgrA and AgrC, two proteins that transduce the QS signal. Specifically, AgrC is a membrane kinase that recognizes extracellular AIPs (QS molecule of S. aureus). After AgrC detects the QS signal, AgrA will be subsequently phosphorylated by it, gaining the ability to activate the transcription of genes downstream promoter P2.

Release circuit

The circuit allows the engineered bacteria to express the self-lysing enzyme complex (Spn1s_LysRZ) under the presence of S. aureus QS signal. Specifically, phosphorylated AgrA could activate the transcription activity of the P2 promoter, enabling the Spn1s_LysRZ located downstream to be transcribed.

For detailed characterization of Spn1s_LysRZ, see BBa_K4593001

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 439
    Illegal BglII site found at 6064
    Illegal BamHI site found at 1323
    Illegal XhoI site found at 2618
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 5716
    Illegal NgoMIV site found at 6548
    Illegal NgoMIV site found at 7222
    Illegal AgeI site found at 3117
    Illegal AgeI site found at 3255
    Illegal AgeI site found at 5585
    Illegal AgeI site found at 6692
    Illegal AgeI site found at 6920
    Illegal AgeI site found at 7091
    Illegal AgeI site found at 7204
  • 1000
    COMPATIBLE WITH RFC[1000]


[edit]
Categories
Parameters
None