Protein interaction networks as starting points to identify novel antimicrobial drug targets

Current Opinion in Microbiology, Volume 16: 566-572, 2013
Roya Zoraghi, Neil E Reiner

Novel classes of antimicrobials are needed to address the challenge of multidrug-resistant bacteria. Current bacterial drug targets mainly consist of specific proteins or subsets of proteins without regard for either how these targets are integrated in cellular networks or how they may interact with host proteins. However, proteins rarely act in isolation, and the majority of biological processes are dependent on interactions with other proteins. Consequently, protein-protein interaction (PPI) networks offer a realm of unexplored potential for next-generation drug targets. In this review, we argue that the architecture of bacterial or host-pathogen protein interactomes can provide invaluable insights for the identification of novel antibacterial drug targets. {PDF}

Assembly of the Eukaryotic PLP-Synthase Complex fromPlasmodium and Activation of the Pdx1 Enzyme

Structure, Volume 20: 172-184, 2012
Gabriela Guédez, Katharina Hipp, Volker Windeisen, Bianca Derrer, Martin Gengenbacher, Bettina Böttcher, Irmgard Sinning, Barbara Kappes, Ivo Tews

Biosynthesis of vitamins is fundamental to malaria parasites. Plasmodia synthesize the active form of vitamin B6 (pyridoxal 5′-phosphate, PLP) using a PLP synthase complex. The EM analysis shown here reveals a random association pattern of up to 12 Pdx2 glutaminase subunits to the dodecameric Pdx1 core complex. Interestingly, Plasmodium falciparum PLP synthase organizes in fibers. The crystal structure shows differences in complex formation to bacterial orthologs as interface variations. Alternative positioning of an α helix distinguishes an open conformation from a closed state when the enzyme binds substrate. The pentose substrate is covalently attached through its C1 and forms a Schiff base with Lys84. Ammonia transfer between Pdx2 glutaminase and Pdx1 active sites is regulated by a transient tunnel. The mutagenesis analysis allows defining the requirement for conservation of critical methionines, whereas there is also plasticity in ammonia tunnel construction as seen from comparison across different species. {PDF}

The Cell as a Collection of Protein Machines: Preparing the Next Generation of Molecular Biologists

Cell, Volume 92: 291–294, 1998
Bruce Alberts

"... It is my hope that some of the young scientists who read this issue of Cell will come to the realization that much of the great future in biology lies in gaining a detailed understanding of the inner workings of the cell's many marvelous protein machines. With this perspective, students may well be motivated to gain the background in the quantitative sciences that they will need to explore this subject successfully. But they will need the faculty in our colleges and universities to lead them." {PDF}

How crowded is the cytoplasm?

Cell, Volume 30: 345-347, 1982
Alice B. Fulton

"... These different strands of evidence support a model of the cytoplasm that is compact and only a few times more open then a crystal. This compact cytoplasm has long been reflected in the dense and complex
images seen in electron micrographs. The complex latticework with persistent elements in it is held together partly by high-affinity interactions and partly by the nonideal behavior of proteins under physiological conditions. The latticework itself is thinly coated with a phase of water that behaves unlike bulk water, but that nevertheless may be the phase of water most important in normal metabolic activities. In the cytoplasm, some proteins clearly exist free in solution. In many cells, however, that solution is not adequately modeled by a dilute solution of proteins. Equally clearly, some proteins are more reasonably described as participating in a crystal, an extensive, three-dimensional, persistent structure in which the relative position of the proteins is maintained. The challenge that lies ahead is to frame more experiments that determine how particular proteins behave in the highly “nonideal” interior of the cell." {PDF}

Fine-Tuning Multiprotein Complexes Using Small Molecules

ACS Chemical Biology, Volume 7: 1311-1320, 2012
Andrea D. Thompson, Amanda Dugan, Jason E. Gestwicki, and Anna K. Mapp

Multiprotein complexes such as the transcriptional machinery, signaling hubs, and protein folding machines are typically composed of at least one enzyme combined with multiple non-enzymes. Often the components of these complexes are incorporated in a combinatorial manner, in which the ultimate composition of the system helps dictate the type, location, or duration of cellular activities. Although drugs and chemical probes have traditionally targeted the enzyme components, emerging strategies call for controlling the function of protein complexes by modulation of protein–protein interactions (PPIs). However, the challenges of targeting PPIs have been well documented, and the diversity of PPIs makes a “one-size-fits-all” solution highly unlikely. These hurdles are particularly daunting for PPIs that encompass large buried surface areas and those with weak affinities. In this Review, we discuss lessons from natural systems, in which allostery and other mechanisms are used to overcome the challenge of regulating the most difficult PPIs. These systems may provide a blueprint for identifying small molecules that target challenging PPIs and affecting molecular decision-making within multiprotein systems. {PDF}

Host Proteins Involved in HIV Infection: New Therapeutic Targets

Biochimica et Biophysica Acta, Volume 1802: 313-321, 2010
Nathalie Arhel and Frank Kirchhoff

ABSTRACT: Current treatment of HIV/AIDS consists of a combination of three to five agents targeting different viral proteins, i.e. the reverse transcriptase, protease, integrase and envelope, and aims to suppress viral replication below detectable levels. This “highly active antiretroviral therapy” (HAART) has brought an enormous benefit for life expectancy and quality in HIV-1-infected individuals, at least in industrialized countries. However, significant limitations with regard to efficiency, drug resistance, side effect and costs still exist. Recent data suggest that cellular factors also represent useful targets for therapy. Here, we summarize findings from several genome-wide screens that identified a large number of cellular factors exploited by HIV-1 at each step of its life cycle. Furthermore, we discuss the evidence that humans are equipped with powerful intrinsic defense mechanisms against retroviruses but that HIV-1 has evolved elaborate ways to counteract or evade them. Preventing the use of host cell proteins obligatory for viral replication or strengthening the cellular defense mechanisms may help to reduce viral replication to harmless levels. A better understanding of the host factors that promote or restrict HIV-1 replication may thus lead to the development of novel therapeutics against HIV/AIDS.

A Physical and Regulatory Map of Host-Influenza Interactions Reveals Pathways in H1N1 Infection

Cell, Volume 139: 1255-1267, 2009
Sagi D. Shapira, Irit Gat-Viks, Bennett O.V. Shum, Amelie Dricot, Marciela M. de Grace, Liguo Wu, Piyush B. Gupta, Tong Hao, Serena J. Silver, David E. Root, David E. Hill, Aviv Regev, and Nir Hacohen

ABSTRACT: During the course of a viral infection, viral proteins interact with an array of host proteins and pathways. Here, we present a systematic strategy to elucidate the dynamic interactions between H1N1 influenza and its human host. A combination of yeast two-hybrid analysis and genome-wide expression profiling implicated hundreds of human factors in mediating viral-host interactions. These factors were then examined functionally through depletion analyses in primary lung cells. The resulting data point to potential roles for some unanticipated host and viral proteins in viral infection and the host response, including a network of RNA-binding proteins, components of WNT signaling, and viral polymerase subunits. This multilayered approach provides a comprehensive and unbiased physical and regulatory model of influenza-host interactions and demonstrates a general strategy for uncovering complex host-pathogen relationships.

Human host factors required for influenza virus replication.

Nature, Volume 463: 813-817, 2010
König R, Stertz S, Zhou Y, Inoue A, Hoffmann HH, Bhattacharyya S, Alamares JG, Tscherne DM, Ortigoza MB, Liang Y, Gao Q, Andrews SE, Bandyopadhyay S, De Jesus P, Tu BP, Pache L, Shih C, Orth A, Bonamy G, Miraglia L, Ideker T, García-Sastre A, Young JA, Palese P, Shaw ML, Chanda SK.

ABSTRACT: Influenza A virus is an RNA virus that encodes up to 11 proteins and this small coding capacity demands that the virus use the host cellular machinery for many aspects of its life cycle1. Knowledge of these host cell requirements not only informs us of the molecular pathways exploited by the virus but also provides further targets that could be pursued for antiviral drug development. Here we use an integrative systems approach, based on genome-wide RNA interference screening, to identify 295 cellular cofactors required for early-stage influenza virus replication. Within this group, those involved in kinase-regulated signalling, ubiquitination and phosphatase activity are the most highly enriched, and 181 factors assemble into a highly significant host–pathogen interaction network. Moreover, 219 of the 295 factors were confirmed to be required for efficient wild-type influenza virus growth, and further analysis of a subset of genes showed 23 factors necessary for viral entry, including members of the vacuolar ATPase (vATPase) and COPI-protein families, fibroblast growth factor receptor (FGFR) proteins, and glycogen synthase kinase 3 (GSK3)-β. Furthermore, 10 proteins were confirmed to be involved in post-entry steps of influenza virus replication. These include nuclear import components, proteases, and the calcium/calmodulin-dependent protein kinase (CaM kinase) IIβ (CAMK2B). Notably, growth of swine-origin H1N1 influenza virus is also dependent on the identified host factors, and we show that small molecule inhibitors of several factors, including vATPase and CAMK2B, antagonize influenza virus replication.