During this year, we have designed and characterized 33 parts in total, containing 10 new basic parts, 5 new composite parts and 7 new improved parts.

Soft robots are easily damaged during ocean exploration. However, it's hard and expensive to repair in time. Aiming to solve this issue, our project designed a new type of material for both self-healing and adhering through tandem repeat polypeptides with n repetitions (TRn). We connected TRn4 with mfp5 to create a "double-sided adhesive", thus attaching squid ring proteins (BBa_K5398001) with different repeat numbers to the substrate. Tyrosine residues on the surface of the Mfp5 could be catalyzed into L-DOPA in the presence of tyrosinase (TyrVs, BBa_K5398600). TRn4 could connect with other highly repetitive tandem repetitions (TRn) of squid ring teeth proteins through the common β-sheet. Additionally, due to the special sequences in the squid teeth ring protein, we employed the self-cleaving RNA cyclase ribozyme (BBa_K5398002 and BBa_K5398003) to design a circular mRNA which could repeatedly translate the sequence and produce the proteins with different repeat numbers.

We showed how we designed and tested the parts in Design and documented them in part registry. Please click the names for more details.

Table 1 | Total Parts

This year, NAU-CHINA designed 10 new basic parts to produce a new type of materials with both self-healing and adhensive capabilities for soft robots and resolve the leakage issue of the BBa_K3739064 at the level of transcription in our improvement project.

TRn derived from squid ring teeth proteins (BBa_K5398001) exhibit strong self-healing abilities.

During transcription of the TRn, the intro spontaneously connects the the 5' intron of td gene from T4 phage (BBa_K5398002) and the 3' intron of td gene from T4 phage (BBa_K5398003) through the catalytic action of guanosine, thus forming a back-splice junction which links the exons into a circular structure.

Aiming to adhere the self-healing materials to the surface of soft robots, the Mfp5 protein, which is rich in tyrosine residues, could be catalyzed into L-DOPA through tyrosinase TyrVs (BBa_K5398600). To prevent the issue of over-oxidation, we combined Mfp6 (BBa_K5398601) with this system.

LicT^CAT-ViViD fusion protein (BBa_K5398611) is a fusion protein which can be activated with blue light. When induced by blue light, LicV undergoes structural alterations and attaches to the ribonucleic antiterminator (RAT) RNA sequence (BBa_K5398612), effectively inhibiting the formation of an RNA terminator stem–loop structure.

Table 1 | Basic Parts

To optimize the system and better implement our expected design, we designed 7 new composite components.

We designed the TRn4-mfp5 (BBa_K5398030) which acts as a "double-sided adhesive" to adhere the self-healing materials to the surface. Then we verified the feasibility and function of combinating the TyrVs with Mfp6 by constructing pET-SUMO-TyrVs (BBa_K5398610) and pET-28a(+)-Mfp6 (BBa_K5398605). In order to contrast the function of cmRNA and TRn5, we constructed pET29a(+)-TRn5 (BBa_K5398005) and pET29a(+)-cmRNA (BBa_K5398006) and characterize them. The detailed results are provided on the registry page.

In the improved part, we constructed the pQE-60-LicV (BBa_K5398630) and pCDFDuet-1-EL222-RAT (BBa_K5398631) to verify the feasibility of coexpression.

Table 1 | Composite Parts