An effervescent exploration of the global history and myriad symbolic meanings of carbonated beverages. More than eighty years before the invention of Coca-Cola, sweet carbonated drinks became ...popular around the world, provoking arguments remarkably similar to those they prompt today. Are they medicinally, morally, culturally, or nutritionally good or bad? Seemingly since their invention, they have been loved-and hated-for being cold or sweet or fizzy or stimulating. Many of their flavors are international: lemon and ginger were more popular than cola until about 1920. Some are local: tarragon in Russia, cucumber in New York, red bean in Japan, and chinotto (exceedingly bitter orange) in Italy. This book looks not only at how something made from water, sugar, and soda became big business, but also how it became deeply important to people-for fizzy drinks' symbolic meanings are far more complex than the water, gas, and sugar from which they are made.
Show your cap or be packaged into HIV-1 Levin, Judith G; Rein, Alan
Proceedings of the National Academy of Sciences - PNAS,
10/2021, Letnik:
118, Številka:
40
Journal Article
The HIV -1 nucleocapsid protein (NC) is a nucleic acid chaperone, which remodels nucleic acid structures so that the most thermodynamically stable conformations are formed. This activity is essential ...for virus replication and has a critical role in mediating highly specific and efficient reverse transcription. NC's function in this process depends upon three properties: (1) ability to aggregate nucleic acids; (2) moderate duplex destabilization activity; and (3) rapid on-off binding kinetics. Here, we present a detailed molecular analysis of the individual events that occur during viral DNA synthesis and show how NC's properties are important for almost every step in the pathway. Finally, we also review biological aspects of reverse transcription during infection and the interplay between NC, reverse transcriptase and human APOBEC3G, an HIV-1 restriction factor that inhibits reverse transcription and virus replication in the absence of the HIV-1 Vif protein.
Human APOBEC3B (A3B) is a member of the APOBEC3 (A3) family of cytidine deaminases, which function as DNA mutators and restrict viral pathogens and endogenous retrotransposons. Recently, A3B was ...identified as a major source of genetic heterogeneity in several human cancers. Here, we determined the solution nuclear magnetic resonance structure of the catalytically active C-terminal domain (CTD) of A3B and performed detailed analyses of its deaminase activity. The core of the structure comprises a central five-stranded β-sheet with six surrounding helices, common to all A3 proteins. The structural fold is most similar to that of A3A and A3G-CTD, with the most prominent difference being found in loop 1. The catalytic activity of A3B-CTD is ∼15-fold lower than that of A3A, although both exhibit a similar pH dependence. Interestingly, A3B-CTD with an A3A loop 1 substitution had significantly increased deaminase activity, while a single-residue change (H29R) in A3A loop 1 reduced A3A activity to the level seen with A3B-CTD. This establishes that loop 1 plays an important role in A3-catalyzed deamination by precisely positioning the deamination-targeted C into the active site. Overall, our data provide important insights into the determinants of the activities of individual A3 proteins and facilitate understanding of their biological function.
Human APOBEC3A is a single-stranded DNA cytidine deaminase that restricts viral pathogens and endogenous retrotransposons, and has a role in the innate immune response. Furthermore, its potential to ...act as a genomic DNA mutator has implications for a role in carcinogenesis. A deeper understanding of APOBEC3A's deaminase and nucleic acid-binding properties, which is central to its biological activities, has been limited by the lack of structural information. Here we report the nuclear magnetic resonance solution structure of APOBEC3A and show that the critical interface for interaction with single-stranded DNA substrates includes residues extending beyond the catalytic centre. Importantly, by monitoring deaminase activity in real time, we find that A3A displays similar catalytic activity on APOBEC3A-specific TTCA- or A3G-specific CCCA-containing substrates, involving key determinants immediately 5' of the reactive C. Our results afford novel mechanistic insights into APOBEC3A-mediated deamination and provide the structural basis for further molecular studies.
APOBEC3G (A3G), a host protein that inhibits HIV-1 reverse transcription and replication in the absence of Vif, displays cytidine deaminase and single-stranded (ss) nucleic acid binding activities. ...HIV-1 nucleocapsid protein (NC) also binds nucleic acids and has a unique property, nucleic acid chaperone activity, which is crucial for efficient reverse transcription. Here we report the interplay between A3G, NC and reverse transcriptase (RT) and the effect of highly purified A3G on individual reactions that occur during reverse transcription. We find that A3G did not affect the kinetics of NC-mediated annealing reactions, nor did it inhibit RNase H cleavage. In sharp contrast, A3G significantly inhibited all RT-catalyzed DNA elongation reactions with or without NC. In the case of (−) strong-stop DNA synthesis, the inhibition was independent of A3G's catalytic activity. Fluorescence anisotropy and single molecule DNA stretching analyses indicated that NC has a higher nucleic acid binding affinity than A3G, but more importantly, displays faster association/disassociation kinetics. RT binds to ssDNA with a much lower affinity than either NC or A3G. These data support a novel mechanism for deaminase-independent inhibition of reverse transcription that is determined by critical differences in the nucleic acid binding properties of A3G, NC and RT.
The human APOBEC3 proteins are a family of DNA-editing enzymes that play an important role in the innate immune response against retroviruses and retrotransposons. APOBEC3G is a member of this family ...that inhibits HIV-1 replication in the absence of the viral infectivity factor Vif. Inhibition of HIV replication occurs by both deamination of viral single-stranded DNA and a deamination-independent mechanism. Efficient deamination requires rapid binding to and dissociation from ssDNA. However, a relatively slow dissociation rate is required for the proposed deaminase-independent roadblock mechanism in which APOBEC3G binds the viral template strand and blocks reverse transcriptase-catalysed DNA elongation. Here, we show that APOBEC3G initially binds ssDNA with rapid on-off rates and subsequently converts to a slowly dissociating mode. In contrast, an oligomerization-deficient APOBEC3G mutant did not exhibit a slow off rate. We propose that catalytically active monomers or dimers slowly oligomerize on the viral genome and inhibit reverse transcription.
Human APOBEC3A (A3A) is a single-domain cytidine deaminase that converts deoxycytidine residues to deoxyuridine in single-stranded DNA (ssDNA). It inhibits a wide range of viruses and endogenous ...retroelements such as LINE-1, but it can also edit genomic DNA, which may play a role in carcinogenesis. Here, we extend our recent findings on the NMR structure of A3A and report structural, biochemical and cell-based mutagenesis studies to further characterize A3A's deaminase and nucleic acid binding activities. We find that A3A binds ssRNA, but the RNA and DNA binding interfaces differ and no deamination of ssRNA is detected. Surprisingly, with only one exception (G105A), alanine substitution mutants with changes in residues affected by specific ssDNA binding retain deaminase activity. Furthermore, A3A binds and deaminates ssDNA in a length-dependent manner. Using catalytically active and inactive A3A mutants, we show that the determinants of A3A deaminase activity and anti-LINE-1 activity are not the same. Finally, we demonstrate A3A's potential to mutate genomic DNA during transient strand separation and show that this process could be counteracted by ssDNA binding proteins. Taken together, our studies provide new insights into the molecular properties of A3A and its role in multiple cellular and antiviral functions.
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