scholarly journals A Novel Domain Assembly Routine for Creating Full-Length Models of Membrane Proteins from Known Domain Structures

Biochemistry ◽  
2017 ◽  
Vol 57 (13) ◽  
pp. 1939-1944 ◽  
Author(s):  
Julia Koehler Leman ◽  
Richard Bonneau
Keyword(s):  
Membrane Proteins ◽  
Full Length ◽  
Domain Structures

scholarly journals A novel domain assembly routine for creating full-length models of membrane proteins from known domain structures

2017 ◽  
Author(s):  
Julia Koehler Leman ◽  
Richard Bonneau
Keyword(s):  
Membrane Proteins ◽  
Biological Function ◽  
Transmembrane Domain ◽  
Search Space ◽  
Full Length ◽  
Conformational Search ◽  
Membrane Domains ◽  
Native Structure ◽  
Domain Structures ◽  
Fragment Insertion

AbstractMembrane proteins composed of soluble and membrane domains are often studied one domain at a time. However, to understand the biological function of entire protein systems and their interactions with each other and drugs, knowledge of full-length structures or models is required. Although few computational methods exist that could potentially be used to model full-length constructs of membrane proteins, none of these methods are perfectly suited for the problem at hand. Existing methods either require an interface or knowledge of the relative orientations of the domains, are not designed for domain assembly, and none of them are developed for membrane proteins. Here we describe the first domain assembly protocol specifically designed for membrane proteins that assembles intra- and extracellular soluble domains and the transmembrane domain into models of the full-length membrane protein. Our protocol does not require an interface between the domains and samples possible domain orientations based on backbone dihedrals in the flexible linker regions, created via fragment insertion, while keeping the transmembrane domain fixed in the membrane. Our method, mp_domain_assembly, implemented in RosettaMP samples domain orientations close to the native structure and is best used in conjunction with experimental data to reduce the conformational search space.

scholarly journals Phage display of functional, full-length human and viral membrane proteins

2008 ◽  
Vol 18 (22) ◽  
pp. 5937-5940 ◽  
Author(s):  
Sudipta Majumdar ◽  
Agnes Hajduczki ◽  
Aaron S. Mendez ◽  
Gregory A. Weiss
Keyword(s):  
Membrane Proteins ◽  
Phage Display ◽  
Full Length ◽  
Viral Membrane

scholarly journals Going native: Direct high throughput screening of secreted full-length IgG antibodies against cell membrane proteins

mAbs ◽  
2017 ◽  
Vol 9 (8) ◽  
pp. 1253-1261 ◽  
Author(s):  
Yongliang Fang ◽  
Thach H. Chu ◽  
Margaret E. Ackerman ◽  
Karl E. Griswold
Keyword(s):  
Membrane Proteins ◽  
Cell Membrane ◽  
High Throughput ◽  
High Throughput Screening ◽  
Full Length ◽  
Igg Antibodies ◽  
Cell Membrane Proteins ◽  
Going Native

scholarly journals Uniformly Aligned Full-Length Membrane Proteins in Liquid Crystalline Bilayers for Structural Characterization

2007 ◽  
Vol 129 (17) ◽  
pp. 5304-5305 ◽  
Author(s):  
Conggang Li ◽  
Philip Gao ◽  
Huajun Qin ◽  
Rose Chase ◽  
Peter L. Gor'kov ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Structural Characterization ◽  
Liquid Crystalline ◽  
Full Length

scholarly journals Direct reconstitution and study of SUN protein interactions in vitro using mammalian cell-free expression

2021 ◽  
Author(s):  
Sagardip Majumder ◽  
Yen-Yu Hsu ◽  
Allen P Liu
Keyword(s):  
Membrane Proteins ◽  
Mammalian Cell ◽  
Protein Function ◽  
Mammalian Cells ◽  
Expression System ◽  
Full Length ◽  
Free Expression ◽  
Lipid Bilayer Membranes ◽  
Linc Complex

SUN proteins are an integral part of LINC (Linker of Nucleoskeleton and Cytoskeleton) complex which spans the nuclear envelope and acts as a physical tether between the cytoskeletal filaments and the nuclear lamina. Several human diseases associated with nuclear deformation are primarily caused by impaired functioning of SUN proteins. Studies in yeast and mammalian cells have illustrated the detrimental effects of different SUN mutants in nuclear positioning and movement. While cell-based studies provide physiological relevance to the functioning of a protein, in vitro reconstitution of isolated proteins is useful in mechanistically dissecting protein function in a biochemically defined environment. In this study, we used a mammalian cell-free expression system to synthesize and reconstitute SUN proteins in artificial lipid bilayer membranes. Building on our previous work demonstrating directional reconstitution of full-length SUN proteins, we deciphered the mechanism of such protein reconstitution and leveraged it to test several theories/models of LINC complex assembly. By using a simple fluorescence-based assay, we revealed the importance of cations such as calcium and the presence of disulfide bonds in the formation of LINC complexes. Through sequential reconstitutions of SUN proteins and soluble luminal domains of SUN proteins, we found that coiled coil domains of SUN proteins are necessary for homomeric and heteromeric interactions of reconstituted SUN proteins. Overall, our results provide mechanistic insights on LINC complex formation and how this might impact cellular mechanotransduction. The facile approach for reconstituting full-length membrane proteins can be extended to study other difficult-to-study membrane proteins in vitro.

scholarly journals The Class I Antigen-processing Pathway for the Membrane Protein Tyrosinase Involves Translation in the Endoplasmic Reticulum and Processing in the Cytosol

1998 ◽  
Vol 187 (1) ◽  
pp. 37-48 ◽  
Author(s):  
Claudio A. Mosse ◽  
Leslie Meadows ◽  
Chance J. Luckey ◽  
David J. Kittlesen ◽  
Eric L. Huczko ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Major Histocompatibility Complex ◽  
Membrane Proteins ◽  
Membrane Protein ◽  
Major Histocompatibility Complex Class ◽  
Antigen Processing ◽  
Full Length ◽  
Class I ◽  
Complex Class ◽  
Histocompatibility Complex

Formation of major histocompatibility complex class I–associated peptides from membrane proteins has not been thoroughly investigated. We examined the processing of an HLA-A*0201–associated epitope, YMDGTMSQV, that is derived from the membrane protein tyrosinase by posttranslational conversion of the sequence YMNGTMSQV. Only YMDGTMSQV and not YMNGTMSQV was presented by HLA-A*0201 on cells expressing full-length tyrosinase, although both peptides have similar affinities for HLA-A*0201 and are transported by TAP. In contrast, translation of YMNGTMSQV in the cytosol, as a minigene or a larger fragment of tyrosinase, led to the presentation of the unconverted YMNGTMSQV. This was not due to overexpression leading to saturation of the processing/conversion machinery, since presentation of the converted peptide, YMDGTMSQV, was low or undetectable. Thus, presentation of unconverted peptide was associated with translation in the cytosol, suggesting that processing of the full-length tyrosinase occurs after translation in the endoplasmic reticulum. Nevertheless, presentation of YMDGTMSQV in cells expressing full-length tyrosinase was TAP (transporter associated with antigen processing) and proteasome dependent. After inhibition of proteasome activity, tyrosinase species could be detected in the cytosol. We propose that processing of tyrosinase involves translation in the endoplasmic reticulum, export of full-length tyrosinase to the cytosol, and retransport of converted peptides by TAP for association with HLA-A*0201.

Proteomics on Full-Length Membrane Proteins Using Mass Spectrometry†

Biochemistry ◽  
2000 ◽  
Vol 39 (15) ◽  
pp. 4237-4242 ◽  
Author(s):  
Johannes le Coutre ◽  
Julian P. Whitelegge ◽  
Adrian Gross ◽  
Eric Turk ◽  
Ernest M. Wright ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Membrane Proteins ◽  
Full Length
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