Precise mapping of subunits in multiprotein complexes by a versatile electron microscopy label.
Positional knowledge of subunits within multiprotein assemblies is crucial for understanding their function. The topological analysis of protein complexes by electron microscopy has undergone impressive development, but analysis of the exact positioning of single subunits has lagged behind. Here we have developed a clonable approximately 80-residue tag that, upon attachment to a target protein, can recruit a structurally prominent electron microscopy label in vitro. This tag is readily visible on single particles and becomes exceptionally distinct after image processing and classification. Thus, our method is applicable for the exact topological mapping of subunits in macromolecular complexes.
Biochemie-Zentrum der Universität Heidelberg, Heidelberg, Germany.
Dirk
D
Flemming
Karsten
K
Thierbach
Philipp
P
Stelter
Bettina
B
Böttcher
Ed
E
Hurt
D000595:Amino Acid Sequence; D004398:Dyneins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D056827:Endosomal Sorting Complexes Required for Transport / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron / Q000379:methods*; D008969:Molecular Sequence Data; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000235:genetics / Q000648:ultrastructure*; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure; D010449:Peptide Mapping / Q000379:methods*; D021122:Protein Subunits / Q000737:chemistry* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000648:ultrastructure
C511158:DID2 protein, S cerevisiae; D056827:Endosomal Sorting Complexes Required for Transport; D046912:Multiprotein Complexes; C097740:NUP120 protein, S cerevisiae; C097894:NUP84 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D021122:Protein Subunits; C083890:SNF7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D004398:Dyneins