一句短语概括整个页面,不要用overview这类无实际意义的词或词组
页面摘要,或者是单项奖核心点 Our project aims at converting engineered Our project aims at converting engineered Our project aims at converting engineered Our project aims at converting engi neered Our project aims at converting engineered Our project aims at converting engineered Our project aims at converting engineered Our project aims at converting engineered
Parts Table
本页面特殊的 groupparts 自动生成表格,需要先关掉html再打开
<groupparts>iGEM20 Fudan</groupparts>Basic Part
CaAP(Calcium Absorption Peptide) is an artificially designed peptide to promote calcium absorption of intestinal epithelial cells. The sequence design of CaAP is based on calcium-binding peptides, a class of oligopeptides capable of chelating calcium, which have been found in hen egg yolk, cow milk casein, whey, soy, wheat germ and tilapia fish, etc. and are capable of facilitating calcium uptake in Caco-2 cells. Moreover, the biosafety of the peptides used in our design can be guaranteed as the MTT assay have shown their absence of cytotoxicity[1].
The whole sequence of CaAP contains 5 different calcium-binding peptides: GPAGPHGPVG, FDHIVY, YQEPVIAPKL[1], NDEELNK[2], and DHTKE[3], Among these peptides, GPAGPHGPVG, FDHIVY, YQEPVIAPKL and NDEELNK function by forming calcium-peptide complex through Asp and Glu residues[1][2]. As for DHTKE, it interacts with the bivalent calcium ion by carboxyl oxygen and amino nitrogen of Asp and Glu as well as the imidazole nitrogen atoms of His residue[3].
To efficiently express CaAP in the engineered strains(E.coli) and allow them to play a role when secreted into the extracellular space, sequences of the 5 mentioned oligopeptides are linked together through FR junctions(Phe-Arg). Hence, they could be expressed in the form of a whole peptide in the engineered bacteria and later cleaved into functional oligopeptides in the presence of digestive enzymes in the intestinal lumen.
We also added 6x-His tag to the C terminal of CaAP for its purification and characterization.
Composite Part
Besides successful expression in the engineered strains(E.coli), the secretion of CaAP from bacteria to intestine is of vital importance as well. Five signal peptides are selected to mediate CaAP translocation through the bacterial inner membrane, including NSP4, OmpA, DsbA, PelB, PhoA. They are all composed of a N-domain of about 2-10 amino acids, a hydrophobic H-domain of about 10–20 amino acids and a C-domain allowing for the cleavage of the signal peptidase after secretion. By adding a signal peptide signal peptide to the N terminus of CaAP,CaAP can be eventually secreted into the lumen of digestive tract and display its robust function of aiding calcium absorption.
The 5 signal peptides are listed below.
Signal Peptide | Number for Registry |
---|---|
NSP4 | BBa_K3606042 |
OmpA | BBa_K3606043 |
DsbA | BBa_K3606030 |
PelB | BBa_K208004 |
PhoA | BBa_K808028 |
References
[1] Three Newly Isolated Calcium-Chelating Peptides from Tilapia Bone Collagen Hydrolysate Enhance Calcium Absorption Activity in Intestinal Caco-2 Cells. Wanwen Liao, Hui Chen, Wengang Jin, Zhennai Yang, Yong Cao, and Jianyin Miao. Journal of Agricultural and Food Chemistry 2020 68 (7), 2091-2098.
[2] In vitro digestion profile and calcium absorption studies of a sea cucumber ovum derived heptapeptide–calcium complex. Pengbo Cui, Songyi Lin, Ziqi Jin, Beiwei Zhu, Liang Song and Na Sun. Food Funct., 2018,9, 4582-4592.
[3] An Exploration of the Calcium-Binding Mode of Egg White Peptide, Asp-His-Thr-Lys-Glu, and In Vitro Calcium Absorption Studies of Peptide–Calcium Complex. Na Sun, Ziqi Jin, Dongmei Li, Hongjie Yin, and Songyi Lin. Journal of Agricultural and Food Chemistry 2017 65 (44), 9782-9789.