Difference between revisions of "Part:BBa K4390035"

Revision as of 15:33, 13 October 2022

SpyTagged His-tagged M. edulis MT

This part is not compatible with BioBrick RFC10 assembly but is compatible with the iGEM Type IIS Part standard which is also accepted by iGEM.

Usage and Biology

SpyTag is a 13 amino acid-long protein that spontaneously reacts with the SpyCatcher protein (12.3 kDa). Together they form an intermolecular isopeptide bond. This provides an alternative to His-tag purification techniques. Insertion of either the SpyTag or the SpyCatcher sequence into a recombinant protein will allow its purification.

Metallothionein (MT) is a small protein (around 6-7 kDa) which is rich in cysteine. These thiol group in cysteines provide ability to chelate almost all heavy metal ions including Cd2+, Hg2+, Pb2+ and As3+, but had been shown that has higher binding affinity with Hg2+ (Manceau, A. et al., 2019). The ability of chelating heavy metals provides the metal tolerance for its hosts. This MT sequence was obtained from Mytilus edulis, a blue mussel which originally live in aqueous environment (MACKAY E. A. et al.,1993).

This construct is used to express SpyTagged Mytilus edulis Metallothionein, which is used to form Spytag-Spycatcher hydrogel with MT fused.

Sequence and Features

Assembly Compatibility:

10
INCOMPATIBLE WITH RFC[10]
Illegal PstI site found at 212
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 11
Illegal NheI site found at 34
Illegal PstI site found at 212
21
COMPATIBLE WITH RFC[21]
23
INCOMPATIBLE WITH RFC[23]
Illegal PstI site found at 212
25
INCOMPATIBLE WITH RFC[25]
Illegal PstI site found at 212
Illegal NgoMIV site found at 161
1000
COMPATIBLE WITH RFC[1000]

Reference

MACKAY, E. A. et al. (1993) Complete amino acid sequences of five dimeric and four monomeric forms of metallothionein from the edible mussel Mytilus edulis. European journal of biochemistry. 218 (1), 183–194.

Manceau, A. et al. (2019) Mercury(II) Binding to Metallothionein in Mytilus edulis revealed by High Energy‐Resolution XANES Spectroscopy. Chemistry : a European journal. 25 (4), 997–1009.

@@ Line 5: / Line 5: @@
 '''This part is not compatible with BioBrick RFC10 assembly but is compatible with the iGEM Type IIS Part standard [[Help:Standards/Assembly/Type_IIS|which is also accepted by iGEM.]]'''
-===Usage and Biology===
+==Usage and Biology==
 SpyTag is a 13 amino acid-long protein that spontaneously reacts with the SpyCatcher protein (12.3 kDa). Together they form an intermolecular isopeptide bond. This provides an alternative to His-tag purification techniques. Insertion of either the SpyTag or the SpyCatcher sequence into a recombinant protein will allow its purification.