Polyproteins in structural biology

Graphical abstract
Highlights • Structures have been determined for natural and recombinant polyproteins. • Native HIV Gag polyprotein architecture was revealed by cryo-EM of immature capsids. • Recombinant polyprotein technology has resolved sample preparation bottlenecks. • The high-resolution structure of influenza polymerase has been solved. • Single-molecule analysis of polyproteins revealed their folding characteristics.
Polyproteins are chains of covalently conjoined smaller proteins that occur in nature as versatile means to organize the proteome of viruses including HIV. During maturation, viral polyproteins are typically cleaved into the constituent proteins with different biological functions by highly specific proteases, and structural analyses at defined stages of this maturation process can provide clues for antiviral intervention strategies. Recombinant polyproteins that use similar mechanisms are emerging as powerful tools for producing hitherto inaccessible protein targets such as the influenza polymerase, for high-resolution structure determination by X-ray crystallography. Conversely, covalent linking of individual protein subunits into single polypeptide chains are exploited to overcome sample preparation bottlenecks. Moreover, synthetic polyproteins provide a promising tool to dissect dynamic folding of polypeptide chains into three-dimensional architectures in single-molecule structure analysis by atomic force microscopy (AFM). The recent use of natural and synthetic polyproteins in structural biology and major achievements are highlighted in this contribution.