Partnership
1.How it all began
When we initially chose to express Lacc6 inside our engineered strain, the degradation rate was inefficient and unsatisfactory. Unable to formulate a suitable Plan B, we sought help and advice from Professor Yu Zekuan at Fudan University. Through this meeting, we were introduced to the cell surface display technique. The prospect of utilizing this technique was very promising in elevating the sulfadiazine(SDZ) degradation rate. We moved on to adopt a new design incorporating the cell surface display technique where laccases were reexpressed to be on the cell's exterior.
Although this new design brought us back on track to achieving our end goal, we encountered new difficulties. The additional components required for the newly designed engineered strain were known, yet we had no idea which anchor protein to use. Surprisingly, we found out we were not alone in facing this issue.
Through a series of coincidences, we connected with two other teams that were also using the cell surface display technique and were facing the exact same problem as we were. The two other teams were LZU-HS-China-A and BFSU-ICUnited.
LZU-HS-China-A focused on converting chitosan from largely consumed shrimp and crabs into usable chitosan oligosaccharides. Implementing the cell surface display technique greatly increased their input to output proportion of chitosan oligosaccharides.
BFSU-ICUnited, on the other hand, focused on developing a novel BPA detector through the use of electrochemical enzyme biosensors. They needed the cell surface display technique to bypass limitations such as enzyme activity loss when expressed intracellularly.
Placed in a similar position, we collectively decided to overcome the challenge together.
2.Searching for the suitable anchor protein
2.1 Anchor Protein Research
The first step we took was to do extensive research about anchor proteins and what makes up a "good" anchor protein in each of our contexts. Together, we came up with a list of necessary properties that constituted a good anchor protein.
The list goes as such:
1.Having signal peptide sequences or transport signals that guide the fusion protein across the cell membrane
2.Can guide the fusion protein to specific sites on the cell surface
3.Having stable anchor elements that can be recognized at specific sites on the cell surface so that the fusion protein can be immobilized on the cell surface
4.Fusion with foreign proteins will not change its stability, and it is uneasily to be broken down by proteases in or outside the cell
Besides researching what constitutes a qualified anchor protein, we also took care to identify the consequences of misusing one.
There are two negative outcomes to selecting an unsuited anchor protein:
1.The structure of anchor protein and passenger protein is easy to affect each other, interferes with their correct folding, and reduces the activity of passenger protein
2.Different transporter proteins will affect the physiological function of the recipient bacteria (growth instability, etc.)
2.2 Finding Potential Candidates
Keeping in mind the research we conducted earlier together, all three teams set off to find potential protein candidates. Each team was responsible for finding a different anchor protein and conducting an analysis of its advantages.
The three proteins found were ice nucleation protein (INP), Lipoprotein, and S-layer protein, respectively done by LZU-HS-China-C, LZU-HS-China-A, and BFSU-ICUnited.
2.2.1 Ice Nucleation Protein
INP is a frequently used cell surface anchor in protein display for Gram-Negative bacteria.
The advantages of using INP we found are as follows:
1.Stably expressed in a large variety of species of host cells
2.Uneasily to be broken down by proteases in or outside the cell
3.High efficiency of displaying on the surface of the host cell
4.Repeating structures can be cut without influencing the functions, which can be advantageous when carrying passenger proteins with larger molecular weights
5.No auxiliary proteins are required
(Schematic of INP)
2.3 The Judgment Call
Confident in the amount of research we have done, all three teams came back together to make a final decision on which anchor protein to use. During our live discussion, each team presented its findings, and, in the end, a detailed comparison of the characteristics was made.
Ultimately, we decided to use INP as the anchor protein for all three of our project designs. INP was suited for a wide range of host cells and was very durable. We communicated our decision to our mentor, who agreed that we had made the right choice.
All three teams benefited greatly from this cooperation. The integration of the cell surface display technique significantly improved each of our project designs. Beholding the capacity of our cooperation, we decided to elevate our collaboration into a long-term partnership.
3 Spreading The Word
3.1 Problem & Solution
As teams in China, we all used the social media platform called WeChat. Each team has an official subscription account to post educational articles pertaining to synthetic biology and their own project. The concept of the subscription accounts was to promote synthetic biology and propagate our iGEM project. Unfortunately, we lacked attention and views on our articles. Despite our team's persistence in making daily posts that are of good quality, the articles lacked attention.
Once again, we found one another and listed some reasonable causes, and came up with corresponding solutions.
The reasons behind low hits are:
1.Educational articles posted about synthetic biology were challenging for the average person to understand or understand enough to be interested.
2.Although we have made many posts, our accounts only existed for a few months, considerably shorter than widely popular accounts.
3.A major source of our views came from sharing with friends and family. Even though each team member has a unique human resources network, it was not enough.
Aiming to combat these limitations, we devised some appropriate solutions:
1.Instead of releasing articles with the bulk of the content dedicated to hardcore experimental or design processes, we shifted towards the redesigned organisms' application and significance to society. By doing so, we hope the reader will find the content more relatable and appreciate the importance of synthetic biology.
2.The solution for challenges 2 & 3 is to promote each other's works. By utilizing the networks of 3 teams, we could reach out to a much wider and more diverse audience. This could mitigate the hits limitations posed by the fact of being young accounts and the restricted audience when promoting as individual teams.
Over the next months, we adjusted our educational posts and followed our plan. Each team sent a reduced version of the two other team's project descriptions and forwarded the links to the official accounts.
3.2 Results
The methodologies we took were successful. The hits for the three team's project-description post was recorded before and after plan execution. All three teams saw a significant rise in the number of views and likes.
4 Synthetic Biology Education
As we approached the end of the iGEM session, we came together for one last time, hoping to end our amazing partnership on a high note. Since the ideology of iGEM is to foster the synthetic biology community, we decided to cooperate in making an additional education presentation. As usual, each team was responsible for different parts of the presentation. LZU-HS-China-A was given the tasks of poster and PowerPoint preparation, and BFSU-ICUnited was responsible for presenting. Our team prepared the background material that was turned into these presentations. Through our collective efforts, the lecture was quickly completed and ready to go.
BFSU-ICUnited first held a lecture session in Beijing in the name of all three teams, and all three teams together followed up with a live presentation on the same content. The lectures were all successful, and it is to our greatest happiness to see so many attend.
5 Final Words
Forming a lasting partnership with LZU-HS-China-A and BFSU-ICUnited has been crucial to our accomplishments in the competition. Together, we found a vital component for our engineered strains, brought our project to more people's attention, and contributed to spreading the awareness of synthetic biology. As we concentrated on different objectives, we always approached the problem by splitting the workload and embodying the economic concept of "specialization." Through such means, great productivity and quality were consistently maintained.
Behind every product of our work were countless hours of meetings and scheduling. It is LZU-HS-China-C's honor and good fortune to be able to work with LZU-HS-China-A and BFSU-ICUnited. We have not only grown but found joy in the presence of your companionship. We would like to take this opportunity to thank both teams in their contributions and we wish them the best in all to come.