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1. Emerging investigator series: characterization of silver and silver nanoparticle interactions with zinc finger peptides

   https://doi.org/10.1039/C9EN00065H  

   Park, Grace; Amaris, Zoe N.; Eiken, Madeline K.; Baumgartner, Karl V.; Johnston, Kathryn A.; Williams, Mari A.; Marckwordt, Jasmine G.; Millstone, Jill E.; Splan, Kathryn E.; Wheeler, Korin E. (January 2019, Environmental Science: Nano)

   In biological systems, chemical and physical transformations of engineered silver nanomaterials (AgENMs) are mediated, in part, by proteins and other biomolecules. Metalloprotein interactions with AgENMs are also central in understanding toxicity, antimicrobial, and resistance mechanisms. Despite their readily available thiolate and amine ligands, zinc finger (ZF) peptides have thus far escaped study in reaction with AgENMs and their Ag( i ) oxidative dissolution product. We report spectroscopic studies that characterize AgENM and Ag( i ) interactions with two ZF peptides that differ in sequence, but not in metal binding ligands: the ZF consensus peptide CP-CCHC and the C-terminal zinc finger domain of HIV-1 nucleocapsid protein p7 (NCp7_C). Both ZF peptides catalyze AgENM (10 and 40 nm, citrate coated) dissolution and agglomeration, two important AgENM transformations that impact bioreactivity. AgENMs and their oxidative dissolution product, Ag( i )(aq), mediate changes to ZF peptide structure and metalation as well. Spectroscopic titrations of Ag( i ) into apo-ZF peptides show an Ag( i )–thiolate charge transfer band, indicative of Ag( i )–ZF binding. Fluorescence studies of the Zn( ii )–NCp_7 complex indicate that the Ag( i ) also effectively competes with the Zn( ii ) to drive Zn( ii ) displacement from the ZFs. Upon interaction with AgENMs, Zn( ii ) bound ZF peptides show a secondary structural change in circular dichroism spectroscopy toward an apo-like structure. The results suggest that Ag( i ) and AgENMs may alter ZF protein function within the cell. 

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2. Evolutionary Analysis of the Zinc Finger and Homeoboxes Family of Proteins Identifies Multiple Conserved Domains and a Common Early Chordate Ancestor

   https://doi.org/10.1093/gbe/evaa039  

   Nail, Alexandra N; Smith, Jeramiah J; Peterson, Martha L; Spear, Brett T; Das, Sabyasachi (March 2020, Genome Biology and Evolution)

   Abstract The Zinc Fingers and Homeoboxes (Zhx) proteins, Zhx1, Zhx2, and Zhx3, comprise a small family of proteins containing two amino-terminal C2–H2 zinc fingers and four or five carboxy-terminal homeodomains. These multiple homeodomains make Zhx proteins unusual because the majority of homeodomain-containing proteins contain a single homeodomain. Studies in cultured cells and mice suggest that Zhx proteins can function as positive or negative transcriptional regulators. Zhx2 regulates numerous hepatic genes, and all three Zhx proteins have been implicated in different cancers. Because Zhx proteins contain multiple predicted homeodomains, are associated with interesting physiological traits, and seem to be only present in the vertebrate lineage, we investigated the evolutionary history of this small family by comparing Zhx homologs from a wide range of chordates. This analysis indicates that the zinc finger motifs and homeodomains are highly similar among all Zhx proteins and also identifies additional Zhx-specific conserved regions, including a 13 amino acid amino-terminal motif that is nearly identical among all gnathostome Zhx proteins. We found single Zhx proteins in the sea lamprey (Petromyzon marinus) and in the nonvertebrate chordates sea squirt (Ciona intestinalis) and lancelet (Branchiostoma floridae); these Zhx proteins are most similar to gnathostome Zhx3. Based on our analyses, we propose that a duplication of the primordial Zhx gene gave rise to Zhx3 and the precursor to Zhx1 and Zhx2. A subsequent tandem duplication of this precursor generated Zhx1 and Zhx2 found in gnathostomes. 

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3. When resurrected ancestors meet the physics of proteins: studying photoreceptors with a virtual time machine

   Hoff, Wouter D (March 2025, Bulletin of the American Physical Society)

   Important gaps of knowledge remain in understanding how the amino acid sequence of a protein encodes its functional properties. We are approaching this question using an evolutionary strategy. Proteins found in nature often occur in protein families consisting of thousands of related proteins that were derived from a common ancestral protein. Both the function and the sequence of these proteins diverged over evolutionary time scales. This phenomenon provides a window into how sequence variation results in functional variation. Computational methods have been reported that use the information present in the variation of the amino acid sequences of proteins belonging to the same protein family to reconstruct the entire evolutionary history that gave rise to the observed sequence variation. We are using gene synthesis methods to resurrect ancient ancestral proteins that gave rise to functionally distinct proteins in present-day proteins, and are applying this approach to bacterial photoreceptor proteins. Biophysical and spectroscopic characterization of these resurrected ancestors yielded proteins with unexpected properties. We will discuss the implications of these observations for understanding aspects of the mechanistic features causing the observed functional variation. 

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4. Protein codes promote selective subcellular compartmentalization

   https://doi.org/10.1126/science.adq2634  

   Kilgore, Henry R; Chinn, Itamar; Mikhael, Peter G; Mitnikov, Ilan; Van_Dongen, Catherine; Zylberberg, Guy; Afeyan, Lena; Banani, Salman F; Wilson-Hawken, Susana; Lee, Tong Ihn; et al (March 2025, Science)

   Cells have evolved mechanisms to distribute ~10 billion protein molecules to subcellular compartments where diverse proteins involved in shared functions must assemble. In this study, we demonstrate that proteins with shared functions share amino acid sequence codes that guide them to compartment destinations. We developed a protein language model, ProtGPS, that predicts with high performance the compartment localization of human proteins excluded from the training set. ProtGPS successfully guided generation of novel protein sequences that selectively assemble in the nucleolus. ProtGPS identified pathological mutations that change this code and lead to altered subcellular localization of proteins. Our results indicate that protein sequences contain not only a folding code but also a previously unrecognized code governing their distribution to diverse subcellular compartments. 

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5. Comparison and evolutionary analysis of patterns controlling liquid-liquid phase separation and prion formation by a fungal protein

   Chernoff, Yury; Grizel, Anastasia; Stevenson, Jonathan; Anee, Ismat J; Paulius, Kristupas (October 2025, Proceedings of Annual Meeting of the Brazilian Biophysical Society)

   Some proteins, including yeast translation termination factor Sup35 (eRF3) are capable of both stress-induced liquid-liquid phase separation (LLPS) and formation of solid fibrous aggregates (amyloids). Fragmentation and propagation of amyloid fibrils generates transmissible (in yeast, heritable) self-perpetuating protein agents, termed prions. Relationships between these processes are still poorly understood. Previous literature data suggested that the ability of Sup35 orthologs to form a prion is sporadically distributed in fungal evolution, and depends on amino acid composition of Sup35 prion domain (PrD), rather than on a evolutionarily variable specific sequence. We have studied two groups of proteins: 1) fungal Sup35 PrDs of various evolutionary origins, and 2) artificially synthesized “scrambled” variants of Saccharomyces cerevisiae Sup35 PrD, that possess identical amino acid composition but different sequences. These proteins were fused to fluorophores and expressed in S. cerevisiae cells. LLPS and amyloid/prion formation were assessed by fluorescence microscopy and biochemical approaches. Amino acid sequences were analyzed by various computational algorithms. Our data indicates that propagation of prion state strongly depends on the evolutionary distance from the host. In contrast, majority of proteins studied are capable of both LLPS and ability to form amyloid fibrils. These capabilities are associated with specific patterns of PrD amino acid distribution, that are broadly conserved among fungi. Notably, PrDs of different sequences differ from each other by their ability to convert from liquid condensates to amyloids, and relationship between these processes is apparently optimized in evolution. Moreover, heterotypic PrDs are can colocalize with each other within liquid condensates and influence amyloid conversion by each other. To conclude, LLPS and amyloid properties depend on specific evolutionarily conserved sequence patterns, indicating possible important biological roles for these processes. These patterns could potentially be used to predict LLPS and prion potential in other sequence contexts. This work was supported by NSF grant 2345660. 

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