Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids
Protein two-dimensional (2D) crystallization on lipid monolayers is a powerful method for structure determination. This method has been extended using the specific and strong interaction between histidine residues (of an overexpressed protein) and Ni2+ ions tethered at the headgroup of synthetic lip...
| Main Authors: | , , , , , , , , , |
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| Format: | Journal article |
| Language: | English |
| Published: |
2002
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| _version_ | 1797078606998929408 |
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| author | Courty, S Lebeau, L Martel, L Lenne, P Balavoine, F Dischert, W Konovalov, O Mioskowski, C Legrand, J Venien-Bryan, C |
| author_facet | Courty, S Lebeau, L Martel, L Lenne, P Balavoine, F Dischert, W Konovalov, O Mioskowski, C Legrand, J Venien-Bryan, C |
| author_sort | Courty, S |
| collection | OXFORD |
| description | Protein two-dimensional (2D) crystallization on lipid monolayers is a powerful method for structure determination. This method has been extended using the specific and strong interaction between histidine residues (of an overexpressed protein) and Ni2+ ions tethered at the headgroup of synthetic lipids. Understanding and then improving the process of adsorption and crystallization of proteins on a lipid monolayer are prerequisites for the production of large and well-ordered crystals of any soluble or membrane His-tagged proteins. These large high-quality arrays are necessary for structural studies at high resolution. We have investigated the steps of adsorption and 2D crystallization of His-HunR using three different lipids: (i) 2-(bis-carboxymethyl-amino)-6-[2-(1,3-di-O-oleyl-glyceroxy)-acetyl-amino] hexanoic acid nickel- (II) (Ni-NTA-DOGA), which has been previously used, and two specifically designed Ni2+-chelating lipids, (ii) Ni-NTA-BB, which has two branched (B) alkyl chains and (iii) Ni-NTA-BF, a nonsymmetrical lipid with one branched (B) and one fluorinated (F) chain. These three lipids, when spread at the air-water interface, exhibit various fluidity properties. The adsorption and crystallization process have been monitored in situ and in real time using a variety of complementary techniques such as ellipsometry, shear rigidity measurements of the monolayer, and Brewster angle microscopy, and we have also developed X-ray reflectivity analysis to investigate the evolution of the electron density profile of the lipid-protein monolayer. Electron microscopy observations of the protein-lipid layers were also performed. We have found that the fluidity of the lipid monolayer has a marked influence on the rates of protein adsorption and crystallization of His-HupR. When Ni-NTA-BB is used to form the monolayer, it accelerates the process of protein adsorption and the protein crystallization is three times faster than when Ni-NTA-DOGA is used. |
| first_indexed | 2024-03-07T00:34:16Z |
| format | Journal article |
| id | oxford-uuid:80d982c9-b65a-41f0-a754-b657cec31ec1 |
| institution | University of Oxford |
| language | English |
| last_indexed | 2024-03-07T00:34:16Z |
| publishDate | 2002 |
| record_format | dspace |
| spelling | oxford-uuid:80d982c9-b65a-41f0-a754-b657cec31ec12022-03-26T21:26:13ZTwo-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipidsJournal articlehttp://purl.org/coar/resource_type/c_dcae04bcuuid:80d982c9-b65a-41f0-a754-b657cec31ec1EnglishSymplectic Elements at Oxford2002Courty, SLebeau, LMartel, LLenne, PBalavoine, FDischert, WKonovalov, OMioskowski, CLegrand, JVenien-Bryan, CProtein two-dimensional (2D) crystallization on lipid monolayers is a powerful method for structure determination. This method has been extended using the specific and strong interaction between histidine residues (of an overexpressed protein) and Ni2+ ions tethered at the headgroup of synthetic lipids. Understanding and then improving the process of adsorption and crystallization of proteins on a lipid monolayer are prerequisites for the production of large and well-ordered crystals of any soluble or membrane His-tagged proteins. These large high-quality arrays are necessary for structural studies at high resolution. We have investigated the steps of adsorption and 2D crystallization of His-HunR using three different lipids: (i) 2-(bis-carboxymethyl-amino)-6-[2-(1,3-di-O-oleyl-glyceroxy)-acetyl-amino] hexanoic acid nickel- (II) (Ni-NTA-DOGA), which has been previously used, and two specifically designed Ni2+-chelating lipids, (ii) Ni-NTA-BB, which has two branched (B) alkyl chains and (iii) Ni-NTA-BF, a nonsymmetrical lipid with one branched (B) and one fluorinated (F) chain. These three lipids, when spread at the air-water interface, exhibit various fluidity properties. The adsorption and crystallization process have been monitored in situ and in real time using a variety of complementary techniques such as ellipsometry, shear rigidity measurements of the monolayer, and Brewster angle microscopy, and we have also developed X-ray reflectivity analysis to investigate the evolution of the electron density profile of the lipid-protein monolayer. Electron microscopy observations of the protein-lipid layers were also performed. We have found that the fluidity of the lipid monolayer has a marked influence on the rates of protein adsorption and crystallization of His-HupR. When Ni-NTA-BB is used to form the monolayer, it accelerates the process of protein adsorption and the protein crystallization is three times faster than when Ni-NTA-DOGA is used. |
| spellingShingle | Courty, S Lebeau, L Martel, L Lenne, P Balavoine, F Dischert, W Konovalov, O Mioskowski, C Legrand, J Venien-Bryan, C Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title | Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title_full | Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title_fullStr | Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title_full_unstemmed | Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title_short | Two-dimensional crystallization of a histidine-tagged protein on monolayers of fluidity-enhanced Ni2+-chelating lipids |
| title_sort | two dimensional crystallization of a histidine tagged protein on monolayers of fluidity enhanced ni2 chelating lipids |
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