Biology subjects

Waltmann, C.

Publications and source records attributed to Waltmann, C..

5 recordsLinked to original sources

bioRxiv2026

HIV-1 Assembly is Assisted by IRSp53 Through an Interaction that Extends Gag Conformation

Budding of the immature virion is an essential part of the HIV-1 life cycle. However, understanding and inhibiting this process has been challenging, as both simulations and theoretical studies suggest that the energy required to bend the membrane is too high for the complete assembly of a spherical viral particle at the plasma membrane without the assistance of cellular host factors. Recent results suggest that the I-BAR protein IRSp53 may be one such host factor, but the molecular mechanism by which the Gag-IRSp53 interaction promotes HIV-1 assembly and budding remains poorly understood. We employ molecular dynamics simulations, starting from AlphaFold models, to show that the SH3 domain of IRSp53 interacts directly with the MACANTD domain of Gag and we experimentally validate this interaction. Our simulations show that the specificity of the interaction with IRSp53 arises from its ability to extend the MACANTD Gag conformation in a manner similar to RNA binding, which is known to activate Gag assembly. We then computationally design HIV-1 Gag mutants that, as confirmed by simulations, do not undergo conformational changes upon interaction with theIRSp53 SH3 domain. Experimentally, these Gag mutants are tested and shown to accumulate in cells. In addition, for these mutants we observe large quantities of Gag assembling at the plasma membrane but exhibiting large budding defects. These results validate the proposed mechanism by which the Gag-IRSp53 interaction promotes completion of HIV-1 budding and suggest new routes for inhibiting viral assembly or budding.

molecular biology↗

bioRxiv2026

HIV-1 Ribonucleoprotein Complex Organization Facilitates Assembly of the Mature HIV-1 Capsid

The encapsulation of the viral RNA condensate inside the capsid is a crucial step in the late stages of the HIV-1 replication cycle. While it is agreed that RNA encapsulation depends on RNA-bound integrase (IN), that capsid-bound IN-RNA filaments are present within the capsid, and that the ribonucleoprotein (RNP) complex takes up only a fraction of the capsid volume, the timing of filament formation and ribonucleoprotein (RNP) encapsulation are unclear. Here, we ran virion-scale, coarse grained molecular dynamics simulations to identify how the organization of the RNP and IN in particular affects mature capsid assembly and genome encapsulation. These virion-scale simulations allow for direct comparison between experimental capsid morphologies and those self-assembled in the simulations. It was found, when relatively few tetramers are on the RNP surface, that IN tetramers could facilitate RNA encapsulation without filling a significant fraction of the capsid volume, but do not form a capsid-templated filament. When octameric IN-RNA filaments are modeled, it was found that WT-like capsid morphologies form only when the IN is organized as a single long filament, in agreement with recent experimental suggestions. These capsids are more conical and elongated than the capsids assembled around the RNP with tetrameric IN. Thus, we suggest the IN-RNA filament assembles ahead of the capsid and directs it to its final conical morphology, consistent with the experimental observation of less elongated capsids when IN oligomerization is disrupted.

molecular biology↗

bioRxiv2026

Engineering the Self-assembly of Bacterial Microcompartment Shell Proteins via Charged Mutations

AbstractProtein self-assembly is a fundamental biological process of great importance for the design and synthesis of biomaterials. Developing the ability to precisely manipulate protein assembly would greatly expand both our understanding of the process and our biotechnological capabilities. Within bacteria, proteins that self-organize to form bacterial microcompartments (MCPs) offer an excellent model system for studying protein self-assembly and advancing biomaterial design capabilities. MCPs consist of irregular polyhedral shells that encase an enzyme core, functioning as enzymatic nanoreactors. In isolation, the abundant shell proteins of the 1,2-propanediol utilization (Pdu) MCP, PduA and PduJ, have a high propensity to self-assemble into tubular structures, analogous in form to carbon nanotubes. Here, we design and characterize tubular structures formed by hexameric PduA and PduJ proteins. We demonstrate that altering hexamer charge offers a systematic strategy for modulating the higherorder assembly of PduA and PduJ across multiple contexts by integrating molecular dynamics simulations with heterologous overexpression, cell-free, and in vivo Salmonella enterica serovar Typhimurium LT2 experiments. First, using molecular simulation, we find that tube chirality and radius play critical roles in determining structural stability and flexibility. Next, overexpression and cell-free experiments show that increasing the overall negative charge of assembling subunits consistently promotes self-assembly into tubular structures. We find that this holds true in the native MCP system, as these same mutations promote the formation of tubular MCP structures in S. enterica LT2. Our results collectively reveal that both electrostatic interactions and fields generated by charges on proteins can be leveraged to control protein-based nanostructures.

biochemistry↗

bioRxiv2025

Mechanistic Insights into Lenacapavir-Induced Off-Pathway HIV-1 Capsid Assembly

The HIV-1 capsid is a fullerene-like shell composed of hexamer and pentamer arrangements of the capsid (CA) proteins. The cone shape of the capsid is particularly important for packaging the viral genome and coordinating nuclear entry. Lenacapavir (LEN), a potent long-acting inhibitor, has been shown to disrupt capsid morphogenesis by binding at the FG-binding pocket located between neighboring CA subunits. Interestingly, inositol hexakisphosphate (IP6), a cellular polyanion, binds within the central pore of capsid pentamers and some hexamers while playing a key role in regulating the hexamer/pentamer switch. As LEN and IP6 interact with overlapping structural elements, they can compete to influence the capsid assembly pathway and outcomes. Using coarse-grained molecular simulations, we examined capsid assembly across varying IP6 and LEN conditions. Our results reveal a concentration-dependent shift in assembly outcomes: LEN accelerates hexamer assembly and reduces pentamer incorporation, leading to malformed, multilayered, or incomplete capsids. Simulations including a model for the viral ribonucleoprotein (RNP) complex further show that LEN-treated capsids frequently fail to encapsidate the RNA genome, indicating impaired maturation. Our calculations confirm that LEN impairs the formation of high-curvature CA lattice regions necessary for closure, supporting a model of off-pathway assembly as a mechanism of viral inhibition.

biophysics↗

bioRxiv2021

Vertex protein PduN tunes encapsulated pathway performance by dictating bacterial metabolosome morphology

Engineering subcellular organization in microbes shows great promise in addressing bottlenecks in metabolic engineering efforts; however, rules guiding selection of an organization strategy or platform are lacking. Here, we study compartment morphology as a factor in mediating encapsulated pathway performance. Using the 1,2-propanediol utilization microcompartment (Pdu MCP) system from Salmonella enterica serovar Typhimurium LT2, we find that we can shift the morphology of this protein nanoreactor from polyhedral to tubular by removing vertex protein PduN. Analysis of the metabolic function between these Pdu microtubes (MTs) shows that they provide a diffusional barrier capable of shielding the cytosol from a toxic pathway intermediate, similar to native MCPs. However, kinetic modeling suggests that the different surface area to volume ratios of MCP and MT structures alters encapsulated pathway performance. Finally, we report a microscopy-based assay that permits rapid assessment of Pdu MT formation to enable future engineering efforts on these structures.

bioengineering↗