Illuminating protein space with a programmable generative model.

Three billion years of evolution has produced a tremendous diversity of protein molecules, but the full potential of proteins is likely to be much greater. Accessing this potential has been challenging for both computation and experiments because the space of possible protein molecules is much larger than the space of those likely to have functions. Here we introduce Chroma, a generative model for proteins and protein complexes that can directly sample novel protein structures and sequences, and that can be conditioned to steer the generative process towards desired properties and functions.

MqsR is a noncanonical microbial RNase toxin that is inhibited by antitoxin MqsA via steric blockage of substrate binding.

The MqsRA toxin-antitoxin system is a component of the Escherichia coli stress response. Free MqsR, a ribonuclease, cleaves mRNAs containing a 5'-GC-3' sequence causing a global shutdown of translation and the cell to enter a state of dormancy. Despite a general understanding of MqsR function, the molecular mechanism(s) by which MqsR binds and cleaves RNA and how one or more of these activities is inhibited by its cognate antitoxin MqsA is still poorly understood.

Bringing the Heavy Chain to Light: Creating a Symmetric, Bivalent IgG-Like Bispecific.

Bispecific molecules are biologically significant, yet their complex structures pose important manufacturing and pharmacokinetic challenges. Nevertheless, owing to similarities with monoclonal antibodies (mAbs), IgG-like bispecifics conceptually align well with conventional expression and manufacturing platforms and often exhibit potentially favorable drug metabolism and pharmacokinetic (DMPK) properties. However, IgG-like bispecifics do not possess target bivalency and current designs often require tedious engineering and purification to ensure appropriate chain pairing.

A highly potent CD73 biparatopic antibody blocks organization of the enzyme active site through dual mechanisms.

The dimeric ectonucleotidase CD73 catalyzes the hydrolysis of AMP at the cell surface to form adenosine, a potent suppressor of the immune response. Blocking CD73 activity in the tumor microenvironment can have a beneficial effect on tumor eradication and is a promising approach for cancer therapy. Biparatopic antibodies binding different regions of CD73 may be a means to antagonize its enzymatic activity.

Trispecific antibodies enhance the therapeutic efficacy of tumor-directed T cells through T cell receptor co-stimulation.

Despite the significant therapeutic advances provided by immune-checkpoint blockade and chimeric antigen receptor T cell treatments, many malignancies remain unresponsive to immunotherapy. Bispecific antibodies targeting tumor antigens and activating T cell receptor signaling have shown some clinical efficacy; however, providing co-stimulatory signals may improve T cell responses against tumors. Here, we developed a trispecific antibody that interacts with CD38, CD3 and CD28 to enhance both T cell activation and tumor targeting.

Cardiovascular Risk Factors in Airline Pilots.

The health of an airline pilot is imperative to the safe travels of millions of people worldwide. Medical providers evaluate the cardiovascular risks for airline pilots and the medical requirements to obtain and maintain licensure as an airline pilot. It is the role of the occupational health nurse practitioner to evaluate and reduce the risk of cardiovascular disease in this population.

Structure-based engineering to restore high affinity binding of an isoform-selective anti-TGFβ1 antibody.

Metelimumab (CAT192) is a human IgG4 monoclonal antibody developed as a TGFβ1-specific antagonist. It was tested in clinical trials for the treatment of scleroderma but later terminated due to lack of efficacy. Subsequent characterization of CAT192 indicated that its TGFβ1 binding affinity was reduced by ∼50-fold upon conversion from the parental single-chain variable fragment (scFv) to IgG4.

A peculiar IclR family transcription factor regulates para-hydroxybenzoate catabolism in Streptomyces coelicolor.

In Streptomyces coelicolor, we identified a para-hydroxybenzoate (PHB) hydroxylase, encoded by gene pobA (SCO3084), which is responsible for conversion of PHB into PCA (protocatechuic acid), a substrate of the β-ketoadipate pathway which yields intermediates of the Krebs cycle. We also found that the transcription of pobA is induced by PHB and is negatively regulated by the product of SCO3209, which we named PobR. The product of this gene is highly unusual in that it is the apparent fusion of two IclR family transcription factors.

Trispecific broadly neutralizing HIV antibodies mediate potent SHIV protection in macaques.

The development of an effective AIDS vaccine has been challenging because of viral genetic diversity and the difficulty of generating broadly neutralizing antibodies (bnAbs). We engineered trispecific antibodies (Abs) that allow a single molecule to interact with three independent HIV-1 envelope determinants: the CD4 binding site, the membrane-proximal external region (MPER), and the V1V2 glycan site. Trispecific Abs exhibited higher potency and breadth than any previously described single bnAb, showed pharmacokinetics similar to those of human bnAbs, and conferred complete immunity against a mixture of simian-human immunodeficiency viruses (SHIVs) in nonhuman primates, in contrast to single bnAbs.

The MqsR/MqsA toxin/antitoxin system protects Escherichia coli during bile acid stress.

Toxin/antitoxin (TA) systems are ubiquitous within bacterial genomes, and the mechanisms of many TA systems are well characterized. As such, several roles for TA systems have been proposed, such as phage inhibition, gene regulation and persister cell formation. However, the significance of these roles is nebulous due to the subtle influence from individual TA systems.

McbR/YncC: implications for the mechanism of ligand and DNA binding by a bacterial GntR transcriptional regulator involved in biofilm formation.

MqsR-controlled colanic acid and biofilm regulator (McbR, also known as YncC) is the protein product of a highly induced gene in early Escherichia coli biofilm development and has been regarded as an attractive target for blocking biofilm formation. This protein acts as a repressor for genes involved in exopolysaccharide production and an activator for genes involved in stress response. To better understand the role of McbR in governing the switch from exponential growth to the biofilm state, we determined the crystal structure of McbR to 2.

BdcA, a protein important for Escherichia coli biofilm dispersal, is a short-chain dehydrogenase/reductase that binds specifically to NADPH.

The Escherichia coli protein BdcA (previously referred to as YjgI) plays a key role in the dispersal of cells from bacterial biofilms, and its constitutive activation provides an attractive therapeutic target for dismantling these communities. In order to investigate the function of BdcA at a molecular level, we integrated structural and functional studies. Our 2.

Type II toxin/antitoxin MqsR/MqsA controls type V toxin/antitoxin GhoT/GhoS.

Toxin endoribonucleases of toxin/antitoxin (TA) systems regulate protein production by selectively degrading mRNAs but have never been shown to control other TA systems. Here we demonstrate that toxin MqsR of the MqsR/MqsA system enriches toxin ghoT mRNA in vivo and in vitro, since this transcript lacks the primary MqsR cleavage site 5'-GCU. GhoT is a membrane toxin that causes the ghost cell phenotype, and is part of a type V TA system with antitoxin GhoS that cleaves specifically ghoT mRNA.

The Escherichia coli toxin MqsR destabilizes the transcriptional repression complex formed between the antitoxin MqsA and the mqsRA operon promoter.

Bacterial biofilms are complex communities of cells containing an increased prevalence of dormant cells known as persisters, which are characterized by an up-regulation of genes known as toxin-antitoxin (TA) modules. The association of toxins with their cognate antitoxins neutralizes toxin activity, allowing for normal cell growth. Additionally, protein antitoxins bind their own promoters and repress transcription, whereas the toxins serve as co-repressors.

A new type V toxin-antitoxin system where mRNA for toxin GhoT is cleaved by antitoxin GhoS.

Among bacterial toxin-antitoxin systems, to date no antitoxin has been identified that functions by cleaving toxin mRNA. Here we show that YjdO (renamed GhoT) is a membrane lytic peptide that causes ghost cell formation (lysed cells with damaged membranes) and increases persistence (persister cells are tolerant to antibiotics without undergoing genetic change). GhoT is part of a new toxin-antitoxin system with YjdK (renamed GhoS) because in vitro RNA degradation studies, quantitative real-time reverse-transcription PCR and whole-transcriptome studies revealed that GhoS masks GhoT toxicity by cleaving specifically yjdO (ghoT) mRNA.