skip to main content


Title: Ribosomal incorporation of cyclic β-amino acids into peptides using in vitro translation
We demonstrate in vitro incorporation of cyclic β-amino acids into peptides by the ribosome through genetic code reprogramming. Further, we show that incorporation efficiency can be increased through the addition of elongation factor P.  more » « less
Award ID(s):
1716766
NSF-PAR ID:
10188864
Author(s) / Creator(s):
; ; ; ; ;
Date Published:
Journal Name:
Chemical Communications
Volume:
56
Issue:
42
ISSN:
1359-7345
Page Range / eLocation ID:
5597 to 5600
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract

    While several studies have shown that invasive rats can have negative effects on island birds through predation (both direct predation and nest predation), other mechanisms for the effects of invasives on island biota have been given less attention. Here, we explore another potential mechanism by which invasive rats can affect native island birds: the competitive use of common resources. We used stable isotope analyses to estimate the fraction of marine and terrestrial sources incorporated into the tissues of two species of passerines (Troglodytes cobbi, Troglodytidae; andCinclodes antarcticus, Furnariidae) and Norway rats (Rattus norvegicus, Muridae) in the Falkland Islands. These two passerines are absent on islands where rats are present. We found significant incorporation of marine resources in the three species, with the highest incorporation in tissues ofT. cobbi. This species appears to be one of the passerines most reliant on marine sources and the most marine member of the family Troglodytidae. We also used the results of these isotopic analyses to estimate the isotopic niche breadth of each of these species and the isotopic niche overlap among them.Rattus norvegicushad a large isotopic niche that overlapped broadly with those of the two passerine species. We propose that different ways of both depicting and estimating isotopic niche widths are complementary rather than alternative. Our results are consistent with the notion that invasive rats might have an impact on these two species of Falkland Island passerines by using common resources but do not rule out the possibility that part of their effect is through direct predation.

     
    more » « less
  2. Stock, Ann M. (Ed.)
    ABSTRACT Staphylococcus aureus can utilize exogenous fatty acids for phospholipid synthesis. The fatty acid kinase FakA is essential for this utilization by phosphorylating exogenous fatty acids for incorporation into lipids. How FakA impacts the lipid membrane composition is unknown. In this study, we used mass spectrometry to determine the membrane lipid composition and properties of S. aureus in the absence of fakA . We found the fakA mutant to have increased abundance of lipids containing longer acyl chains. Since S. aureus does not synthesize unsaturated fatty acids, we utilized oleic acid (18:1) to track exogenous fatty acid incorporation into lipids. We observed a concentration-dependent incorporation of exogenous fatty acids into the membrane that required FakA. We also tested how FakA and exogenous fatty acids impact membrane-related physiology and identified changes in membrane potential, cellular respiration, and membrane fluidity. To mimic the host environment, we characterized the lipid composition of wild-type and fakA mutant bacteria grown in mouse skin homogenate. We show that wild-type S. aureus can incorporate exogenous unsaturated fatty acids from host tissue, highlighting the importance of FakA in the presence of host skin tissue. In conclusion, FakA is important for maintaining the composition and properties of the phospholipid membrane in the presence of exogenous fatty acids, impacting overall cell physiology. IMPORTANCE Environmental fatty acids can be harvested to supplement endogenous fatty acid synthesis to produce membranes and circumvent fatty acid biosynthesis inhibitors. However, how the inability to use these fatty acids impacts lipids is unclear. Our results reveal lipid composition changes in response to fatty acid addition and when S. aureus is unable to activate fatty acids through FakA. We identify concentration-dependent utilization of oleic acid that, when combined with previous work, provides evidence that fatty acids can serve as a signal to S. aureus . Furthermore, using mouse skin homogenates as a surrogate for in vivo conditions, we showed that S. aureus can incorporate host fatty acids. This study highlights how exogenous fatty acids impact bacterial membrane composition and function. 
    more » « less
  3. null (Ed.)
    Abstract The revolution in understanding higher order chromosome dynamics and organization derives from treating the chromosome as a chain polymer and adapting appropriate polymer-based physical principles. Using basic principles, such as entropic fluctuations and timescales of relaxation of Rouse polymer chains, one can recapitulate the dominant features of chromatin motion observed in vivo. An emerging challenge is to relate the mechanical properties of chromatin to more nuanced organizational principles such as ubiquitous DNA loops. Toward this goal, we introduce a real-time numerical simulation model of a long chain polymer in the presence of histones and condensin, encoding physical principles of chromosome dynamics with coupled histone and condensin sources of transient loop generation. An exact experimental correlate of the model was obtained through analysis of a model-matching fluorescently labeled circular chromosome in live yeast cells. We show that experimentally observed chromosome compaction and variance in compaction are reproduced only with tandem interactions between histone and condensin, not from either individually. The hierarchical loop structures that emerge upon incorporation of histone and condensin activities significantly impact the dynamic and structural properties of chromatin. Moreover, simulations reveal that tandem condensin–histone activity is responsible for higher order chromosomal structures, including recently observed Z-loops. 
    more » « less
  4. β- N -methylamino- l -alanine (BMAA) is a nonproteinogenic amino acid that has been associated with neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD). BMAA has been found in human protein extracts; however, the mechanism by which it enters the proteome is still unclear. It has been suggested that BMAA is misincorporated at serine codons during protein synthesis, but direct evidence of its cotranslational incorporation is currently lacking. Here, using LC-MS–purified BMAA and several biochemical assays, we sought to determine whether any aminoacyl-tRNA synthetase (aaRS) utilizes BMAA as a substrate for aminoacylation. Despite BMAA's previously predicted misincorporation at serine codons, following a screen for amino acid activation in ATP/PP i exchange assays, we observed that BMAA is not a substrate for human seryl-tRNA synthetase (SerRS). Instead, we observed that BMAA is a substrate for human alanyl-tRNA synthetase (AlaRS) and can form BMAA-tRNA Ala by escaping from the intrinsic AlaRS proofreading activity. Furthermore, we found that BMAA inhibits both the cognate amino acid activation and the editing functions of AlaRS. Our results reveal that, in addition to being misincorporated during translation, BMAA may be able to disrupt the integrity of protein synthesis through multiple different mechanisms. 
    more » « less
  5. Abstract

    It is now widely accepted that siderophores play a role in marine iron biogeochemical cycling. However, the mechanisms by which siderophores affect the availability of iron from specific sources and the resulting significance of these processes on iron biogeochemical cycling as a whole have remained largely untested. In this study, we develop a model system for testing the effects of siderophore production on iron bioavailability using the marine copiotroph Alteromonas macleodii ATCC 27126. Through the generation of the knockout cell line ΔasbB::kmr, which lacks siderophore biosynthetic capabilities, we demonstrate that the production of the siderophore petrobactin enables the acquisition of iron from mineral sources and weaker iron-ligand complexes. Notably, the utilization of lithogenic iron, such as that from atmospheric dust, indicates a significant role for siderophores in the incorporation of new iron into marine systems. We have also detected petrobactin, a photoreactive siderophore, directly from seawater in the mid-latitudes of the North Pacific and have identified the biosynthetic pathway for petrobactin in bacterial metagenome-assembled genomes widely distributed across the global ocean. Together, these results improve our mechanistic understanding of the role of siderophore production in iron biogeochemical cycling in the marine environment wherein iron speciation, bioavailability, and residence time can be directly influenced by microbial activities.

     
    more » « less