Search

The initiation of gene transcription by RNA polymerase II is regulated by a plethora of proteins in human cells. The first general transcription factor to bind gene promoters is transcription [...]

The article discusses an automated method for optimizing expression of multiprotein complexes in insect cells by using an advanced microscale bioreactor system. It explains that how the application of laboratory automation can facilitate medium- to high-throughput multicomplex protein expression programs. It outlines various challenges associated with recombinant expression systems for multiprotein complexes.

Background: Due to COVID-19, pandemic preparedness emerges as a key imperative, necessitating new approaches to accelerate development of reagents against infectious pathogens., Methods: Here, we developed an integrated approach combining synthetic, computational and structural methods with in vitro antibody selection and in vivo immunization to design, produce and validate nature-inspired nanoparticle-based reagents against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)., Results: Our approach resulted in two innovations: (i) a thermostable nasal vaccine called ADDoCoV, displaying multiple copies of a SARS-CoV-2 receptor binding motif derived epitope and (ii) a multivalent nanoparticle superbinder, called Gigabody, against SARS-CoV-2 including immune-evasive variants of concern (VOCs). In vitro generated neutralizing nanobodies and electron cryo-microscopy established authenticity and accessibility of epitopes displayed by ADDoCoV. Gigabody comprising multimerized nanobodies prevented SARS-CoV-2 virion attachment with picomolar EC 50 . Vaccinating mice resulted in antibodies cross-reacting with VOCs including Delta and Omicron., Conclusion: Our study elucidates Adenovirus-derived dodecamer (ADDomer)-based nanoparticles for use in active and passive immunization and provides a blueprint for crafting reagents to combat respiratory viral infections., Competing Interests: C.S., K.G. and I.B. report shareholding in Halo Therapeutics Ltd unrelated to this Correspondence. I.B. reports shareholding in Geneva Biotech SARL, unrelated to this correspondence. F.G., J.H. and I.B. report shareholding in Imophoron Ltd, related to this Correspondence. Patents and patent applications have been filed related to ADDomer vaccines and therapeutics (WO2017167988A, EP22191583.8). The other authors do not declare competing interests. ADDomer is a registered trademark of Imophoron Ltd., (© The Author(s) 2023. Published by Oxford University Press on behalf of Antibody Therapeutics. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Baculovirus-based expression of proteins in insect cell cultures has emerged as a powerful technology to produce complex protein biologics for many applications ranging from multiprotein complex structural biology to manufacturing of therapeutic proteins including virus-like particles (VLPs). VLPs are protein assemblies that mimic live viruses but typically do not contain any genetic material, and therefore are safe and attractive alternatives to life attenuated or inactivated viruses for vaccination purposes. MultiBac is an advanced baculovirus expression vector system (BEVS) which consists of an engineered viral genome that can be customized for tailored applications. Here we describe the creation of a MultiBac-based VLP-factory ™ , based on the M1 capsid protein from influenza, and its application to produce in a parallelized fashion an array of influenza-derived VLPs containing functional mutations in influenza hemagglutinin (HA) thought to modulate the immune response elicited by the VLP.

The production of a homogeneous protein sample in sufficient quantities is an essential prerequisite not only for structural investigations but represents also a rate-limiting step for many functional studies. In the cell, a large fraction of eukaryotic proteins exists as large multicomponent assemblies with many subunits, which act in concert to catalyze specific activities. Many of these complexes cannot be obtained from endogenous source material, so recombinant expression and reconstitution are then required to overcome this bottleneck. This chapter describes current strategies and protocols for the efficient production of multiprotein complexes in large quantities and of high quality, using the baculovirus/insect cell expression system.