Difference: TwodxAgrC (11 vs. 12)

Revision 1219 Jul 2013 - Main.NicolasCoudray

 
META TOPICPARENT name="2DCrystallizationExp"

2dx - AgrC 2D-crystallization

Contents

Properties

  • Amino acid sequence:
MELLNSYNFVLFVLTQMILMFTIPAIISGIKYSKLDYFFIIGISTLSLFLFKMFDSASLIILTSFIIIMYFVKIKWYSILLIMTSQIILYCANYMYIVIYAYITKISDSIFVIFPSFFVVYVTISILFSYIINRVLKKISTPYLILNKGFLIVISTILLLTFSLFFFYSQINSDEAKVIRQYSFIFIGITIFLSILTFVISQFLLKEMKYKRNQEEIETYYEYTLKIEAINNEMRKFRHDYVNILTTLSEYIREDDMIGLRAYFNKNIVPMKDNLQMNAIKLNGIENLKVREIKGLITAKILRAQEMNIPISIEIPDEVSSINLNMIDLSRSIGIILDNAIEASTEIDDPIIRVAFIESENSVTFIVMNKCADDIPRIHELFQESFSTKGEGRGLGLSTLKEIADNADNVLLDTIIENGFFIQKVEIINN

  • 430 amino acids
  • 49.8 kDa
  • pI ~ 5.28
  • 44 negatively charged residues
  • 38 positively charged residues
  • extinction coefficient: 38405
  • instability index: 38.13
  • alipathic index: 127.63
  • GRAVY: 0.510
  • TM helices: 6 (Phoebius)
  • amino acids in the TM domain: 138 (LPR(m/m) expected: 5-20 (or 10-160) ~ 0.1 to 0.3 in w/w (or 0.15 to 2.5); with LPR(m/m)=24>>LPR(w/w)=0.4

X216 - Sparse matrix + charged lipids

Protein in 0.14% FC12: X216-Protein_Acquisition_AgrC.doc.

Sparse matrix + charged lipids tried: X216-AgrC.xlsx.

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Results, X216

Aggregates or Aggregates+ vesicles for mostly of the PC lipids
Aggregates/clusters + lots of tiny “textured” vesicles (<150 nm) for most of the E.coli lipids and DOPG mixtures, or a bit with DOPC:DOPA.
Image selection: X216-AgrC-ImageSelection.pptx.
 
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X162

Protein in FC12. DMPC and DOPC in FC12, LPR 0.25 to 1, in 20ul dialysis buttons vs 1 buffer similar to the one in which the protein is (pH8)

Results, X162

Mainly aggregates (also with biobeads tested by Marissa). Protein concentration may be higher than expected (gel may give some info).

Image selection: X162-AgrC.xlsx.

X130CD-X131 - Lipid screening

Protein in 20mM Tris-ClH ph8: 100mM ClNa?: 2mM TCEP: 0.2% DM. Protein acquisition form: June2012-Protein_acquisition_form-ArgC.doc. Conditions:
  • lipids solubilized in DM, like the protein (DMPC, E.coli, DOPG, EggPC?:PA)
  • LPR 0.25 to 1.5
  • pH6 and 7
  • 28 degrees (and a few at 28-37 deg)
  • microdialysis plate (and MBCD robot, reaching Det:CD of 1:3 after 36 hours)
  • buffer for dialysis: 20mM MES, 200mM NaCl, 5mM MgCl2, 5 NaN3?

See details: X130CD-X131-ArgC.xlsx.

Comments, July 2012

Image selection: X130_and_X131_ArgC.ppt. X130CD:
  • lots of vesicles (DMPC)
X131:
  • mostly vesicles are obtained
  • recommended lipid: DMPC (pH 6.0 or 7.0)
  • recommended lipid: DOPG (rather pH 7) look better (big vesicle at LPR 0.5: more images to be acquired)
  • DMPC vesicles appear bigger when cyclodextrin robot has been used.

subsub level topic

a79 2
  • Set ALLOWTOPICVIEW =

-- NicolasCoudray - 19 Jun 2012

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META FILEATTACHMENT attr="" autoattached="1" comment="" date="1340825569" name="X130CD-X131-ArgC.xlsx" path="X130CD-X131-ArgC.xlsx" size="602004" user="Main.NicolasCoudray" version="2"
 
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META FILEATTACHMENT attr="" autoattached="1" comment="" date="1358880868" name="X162-AgrC-matrix.jpg" path="X162-AgrC-matrix.jpg" size="1171566" user="Main.NicolasCoudray" version="1"
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META FILEATTACHMENT attr="" autoattached="1" comment="" date="1355417647" name="X162-AgrC.xlsx" path="X162-AgrC.xlsx" size="608615" user="Main.NicolasCoudray" version="1"
 
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META TOPICMOVED by="NicolasCoudray" date="1390912507" from="Main.TwodxArgC" to="Main.TwodxAgrC"
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