Difference between revisions of "Part:BBa K3889022"
Line 4: Line 4:
  
 
Since ovastacin (<partinfo>BBa_K3889024</partinfo>) has a eukaryotic origin, this protein undergoes several Post Translational Modifications (PTMs) including the addition of disulphide bonds and phosphorylation at serine and tyrosine residues.  
 
Since ovastacin (<partinfo>BBa_K3889024</partinfo>) has a eukaryotic origin, this protein undergoes several Post Translational Modifications (PTMs) including the addition of disulphide bonds and phosphorylation at serine and tyrosine residues.  
−
However, our recombinant protein will be produced in a prokaryotic organism which lacks the necessary genes to carry out PTMs. Hence, we used phosphomimetic variant(s) of ovastacin by changing or mutating the serine to aspartic acid and tyrosine to glutamic acid residues at S99, S156, S244 and Y279 (amino acid numbered with reference to pro-ovastacin) respectively. We then used I-TASSER to predict the structure of this phosphomimetic variant.  
+
However, the recombinant protein if produced in a prokaryotic organism it will lack the necessary genes to carry out PTMs. Hence, a phosphomimetic variant(s) of ovastacin was made by changing or mutating the serine to aspartic acid S99, S156 and S244 (amino acid numbered with reference to pro-ovastacin) respectively.
  
 
===Protein Sequence===
 
===Protein Sequence===
−
RLLSAASNKWPMGGDGVVEVPFLLSSKYDEPSRQVILEALAEFERSTCIRFVTYQDQRDFISIIPMYGCFSDVGRSGGMQVVSLAPTCLQKGRGIVL<strong>HELMHVLGFWH</strong>E HTRADRDRYIRVNWNEILPGFEINFIKSRSSNMLTPYDYSSVMHYGRLAFDRRGLPTITPLWAPSVHIGQRWNLSASDITRVLKLEGCS
+
RLLSAASNKWPMGGDGVVEVPFLLSSKYDEPSRQVILEALAEFERSTCIRFVTYQDQRDFISIIPMYGCFSDVGRSGGMQVVSLAPTCLQKGRGIVL<span style="color:red">HELMHVLGFWH</span>E HTRADRDRYIRVNWNEILPGFEINFIKSRSSNMLTPYDYSSVMHYGRLAFDRRGLPTITPLWAPSVHIGQRWNLSASDITRVLKLYGCS
 +
 
 +
where <span style="color:red">red</span> denotes the active site
 +
<html>
 +
<head>
 +
<style>
 +
div.scroll {overflow-x: aut0; overflow-y: hidden;}
 +
</style>
 +
</head>
 +
<body>
 +
<div class="scroll">
 +
<img src="https://static.igem.org/mediawiki/parts/8/8e/T--IISER-Tirupati_India--ovastacin_allignment.jpg">
 +
</div>
 +
</body>
 +
</html>
  
−
[[File:T--IISER-Tirupati India--Fig3-Phosphomimic Ovastacin.png]]<br>
 
−
Fig 3. Phosphomimetic variant of Ovastacin (aspartic acid residues marked with lime and glutamic acid residue marked with cyan)
 
  
−
{| class=wikitable
 
−
!C-score
 
−
! Estimated TM-score
 
−
! Estimated RMSD
 
−
|-
 
−
| 1.84
 
−
| 0.97±0.05
 
−
| 1.8±1.5Å
 
−
|}
 
  
  

Revision as of 04:56, 18 October 2021


Human Ovastacin protease phosphomimic_A

Since ovastacin (BBa_K3889024) has a eukaryotic origin, this protein undergoes several Post Translational Modifications (PTMs) including the addition of disulphide bonds and phosphorylation at serine and tyrosine residues. However, the recombinant protein if produced in a prokaryotic organism it will lack the necessary genes to carry out PTMs. Hence, a phosphomimetic variant(s) of ovastacin was made by changing or mutating the serine to aspartic acid S99, S156 and S244 (amino acid numbered with reference to pro-ovastacin) respectively.

Protein Sequence

RLLSAASNKWPMGGDGVVEVPFLLSSKYDEPSRQVILEALAEFERSTCIRFVTYQDQRDFISIIPMYGCFSDVGRSGGMQVVSLAPTCLQKGRGIVLHELMHVLGFWHE HTRADRDRYIRVNWNEILPGFEINFIKSRSSNMLTPYDYSSVMHYGRLAFDRRGLPTITPLWAPSVHIGQRWNLSASDITRVLKLYGCS

where red denotes the active site



Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]