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Cold Spring Harbor Perspectives in... Dec 2018This review summarizes our current understanding of the major pathway for the initiation phase of protein synthesis in eukaryotic cells, with a focus on recent advances.... (Review)
Review
This review summarizes our current understanding of the major pathway for the initiation phase of protein synthesis in eukaryotic cells, with a focus on recent advances. We describe the major scanning or messenger RNA (mRNA) mG cap-dependent mechanism, which is a highly coordinated and stepwise regulated process that requires the combined action of at least 12 distinct translation factors with initiator transfer RNA (tRNA), ribosomes, and mRNAs. We limit our review to studies involving either mammalian or budding yeast cells and factors, as these represent the two best-studied experimental systems, and only include a reference to other organisms where particular insight has been gained. We close with a brief description of what we feel are some of the major unknowns in eukaryotic initiation.
Topics: Animals; Eukaryotic Cells; Peptide Chain Initiation, Translational; Protein Biosynthesis
PubMed: 29735639
DOI: 10.1101/cshperspect.a033092 -
Journal of Industrial Microbiology &... Aug 2021Nonribosomal peptide synthetases (NRPS) are large multimodular enzymes that synthesize a diverse variety of peptides. Many of these are currently used as... (Review)
Review
Nonribosomal peptide synthetases (NRPS) are large multimodular enzymes that synthesize a diverse variety of peptides. Many of these are currently used as pharmaceuticals, thanks to their activity as antimicrobials (penicillin, vancomycin, daptomycin, echinocandin), immunosuppressant (cyclosporin) and anticancer compounds (bleomycin). Because of their biotechnological potential, NRPSs have been extensively studied in the past decades. In this review, we provide an overview of the main structural and functional features of these enzymes, and we consider the challenges and prospects of engineering NRPSs for the synthesis of novel compounds. Furthermore, we discuss secondary metabolism and NRP synthesis in the filamentous fungus Penicillium rubens and examine its potential for the production of novel and modified β-lactam antibiotics.
Topics: Penicillium; Peptide Biosynthesis, Nucleic Acid-Independent; Peptide Synthases
PubMed: 34279620
DOI: 10.1093/jimb/kuab045 -
Current Opinion in Chemical Biology Feb 2011Polyketides and non-ribosomal peptides are in a class of natural products important both as drug sources and as dangerous toxins and virulence factors. While studies... (Review)
Review
Polyketides and non-ribosomal peptides are in a class of natural products important both as drug sources and as dangerous toxins and virulence factors. While studies over the last two decades have provided substantial characterization of the modular synthases that produce these compounds at the genetic level, their understanding at the protein level is much less understood. New proteomic platforms called an orthogonal active site identification system (OASIS) and proteomic interrogation of secondary metabolism (PrISM) have been developed to identify and quantify natural product synthase enzymes. Reviewed here, these tools offer the means to discover and analyze modular synthetic pathways that are limited by genetic techniques, opening the tools of contemporary proteomics to natural product sciences.
Topics: Biological Products; Humans; Macrolides; Mass Spectrometry; Peptide Biosynthesis, Nucleic Acid-Independent; Peptides; Proteomics; Ribosomes
PubMed: 21087894
DOI: 10.1016/j.cbpa.2010.10.021 -
Current Opinion in Structural Biology Aug 2013Naturally occurring polyketides and nonribosomal peptides with broad and potent biological activities continue to inspire the discovery of new and improved analogs. The... (Review)
Review
Naturally occurring polyketides and nonribosomal peptides with broad and potent biological activities continue to inspire the discovery of new and improved analogs. The biosynthetic apparatus responsible for the construction of these natural products has been the target of intensive protein engineering efforts. Traditionally, engineering has focused on substituting individual enzymatic domains or entire modules with those of different building block specificity, or by deleting various enzymatic functions, in an attempt to generate analogs. This review highlights strategies based on site-directed mutagenesis of substrate binding pockets, semi-rational mutagenesis, and whole-gene random mutagenesis to engineer the substrate specificity, activity, and protein interactions of polyketide and nonribosomal peptide biosynthetic machinery.
Topics: Antineoplastic Agents; Gene Expression; Mutagenesis, Site-Directed; Peptide Biosynthesis, Nucleic Acid-Independent; Peptide Synthases; Polyketide Synthases; Polyketides; Protein Engineering
PubMed: 23838175
DOI: 10.1016/j.sbi.2013.06.012 -
Biochimica Et Biophysica Acta. Proteins... Nov 2017Nonribosomal peptide synthetases (NRPSs) are incredible macromolecular machines that produce a wide range of biologically- and therapeutically-relevant molecules. During... (Review)
Review
Nonribosomal peptide synthetases (NRPSs) are incredible macromolecular machines that produce a wide range of biologically- and therapeutically-relevant molecules. During synthesis, peptide elongation is performed by the condensation (C) domain, as it catalyzes amide bond formation between the nascent peptide and the amino acid it adds to the chain. Since their discovery more than two decades ago, C domains have been subject to extensive biochemical, bioinformatic, mutagenic, and structural analyses. They are composed of two lobes, each with homology to chloramphenicol acetyltransferase, have two binding sites for their two peptidyl carrier protein-bound ligands, and have an active site with conserved motif HHxxxDG located between the two lobes. This review discusses some of the important insights into the structure, catalytic mechanism, specificity, and gatekeeping functions of C domains revealed since their discovery. In addition, C domains are the archetypal members of the C domain superfamily, which includes several other members that also function as NRPS domains. The other family members can replace the C domain in NRP synthesis, can work in concert with a C domain, or can fulfill diverse and novel functions. These domains include the epimerization (E) domain, the heterocyclization (Cy) domain, the ester-bond forming C domain, the fungal NRPS terminal C domain (C), the β-lactam ring forming C domain, and the X domain. We also discuss structural and function insight into C, E, Cy, C and X domains, to present a holistic overview of historical and current knowledge of the C domain superfamily. This article is part of a Special Issue entitled: Biophysics in Canada, edited by Lewis Kay, John Baenziger, Albert Berghuis and Peter Tieleman.
Topics: Peptide Biosynthesis; Peptide Synthases; Peptides; Protein Domains; Structure-Activity Relationship
PubMed: 28526268
DOI: 10.1016/j.bbapap.2017.05.010 -
The Israel Medical Association Journal... Oct 2012The role of carbon in the development of life and as the structural backbone of all organisms is universally accepted and an essential part of evolution. However, the... (Review)
Review
The role of carbon in the development of life and as the structural backbone of all organisms is universally accepted and an essential part of evolution. However, the molecular basis is largely unknown and the interactions of carbon with nitrogen and oxygen in space are enigmatic. In 1985, the previously unknown form of carbon, coined fullerene, was discovered. We hypothesize that by virtue of the unique properties of fullerene, this hollow, ultra-robust, large, purely carbon molecule was the earliest progenitor of life. It acted as a stable universal biologic template on which small molecules spontaneously assembled and then formed, by further assembly, a surface mantle (here termed rosasome) of larger molecules. We submit that this process, by its inherent flexibility, initiated evolution, allowing the emergence of parallel diverse rosasome lines responding selectively to varying spatial environments. For example, rosasomal lines mantled with nucleotide and peptide layers are conceived as primordial forerunners of the ubiquitous ribosome. Moreover, the parallel independent and interdependent evolution of rosasome lines would be more rapid than sequential development, refute precedence of either DNA or RNA, and explain the evolution of integration of two subunits with different structures and functions in ribosomes and of the triplet nature of the codon. Based on recent astronomical data, this hypothesis supports the concept that life is not a singularity. This concept also suggests a potential vehicle for therapeutics, biotechnology and genetic engineering.
Topics: DNA; Evolution, Molecular; Fullerenes; Humans; Models, Biological; Origin of Life; Peptide Biosynthesis; Ribosomes
PubMed: 23193780
DOI: No ID Found -
Proteins Mar 2022Microviridins, tricyclic peptide natural products originally isolated from cyanobacteria, function as inhibitors of diverse serine-type proteases. Here we report the...
Microviridins, tricyclic peptide natural products originally isolated from cyanobacteria, function as inhibitors of diverse serine-type proteases. Here we report the structure and biochemical characterization of AMdnB, a unique iterative macrocyclase involved in a microviridin biosynthetic pathway from Anabaena sp. PCC 7120. The ATP-dependent cyclase, along with the homologous AMdnC, introduce up to nine macrocyclizations on three distinct core regions of a precursor peptide, AMdnA. The results presented here provide structural and mechanistic insight into the iterative chemistry of AMdnB. In vitro AMdnB-catalyzed cyclization reactions demonstrate the synthesis of the two predicted tricyclic products from a multi-core precursor peptide substrate, consistent with a distributive mode of catalysis. The X-ray structure of AMdnB shows a structural motif common to ATP-grasp cyclases involved in RiPPs biosynthesis. Additionally, comparison with the noniterative MdnB allows insight into the structural basis for the iterative chemistry. Overall, the presented results provide insight into the general mechanism of iterative enzymes in ribosomally synthesized and post-translationally modified peptide biosynthetic pathways.
Topics: Amino Acid Sequence; Benchmarking; Biological Products; Biosynthetic Pathways; Catalysis; Crystallography, X-Ray; Cyanobacteria; Cyclization; Models, Molecular; Peptide Biosynthesis; Peptides, Cyclic; Protein Binding; Protein Conformation; Protein Processing, Post-Translational; Ribosomes
PubMed: 34664307
DOI: 10.1002/prot.26264 -
Nature Communications Jan 2021Nonribosomal peptide synthetases containing starter condensation domains direct the biosynthesis of nonribosomal lipopeptides, which generally exhibit wide...
Nonribosomal peptide synthetases containing starter condensation domains direct the biosynthesis of nonribosomal lipopeptides, which generally exhibit wide bioactivities. The acyl chain has strong impacts on bioactivity and toxicity, but the lack of an in-depth understanding of starter condensation domain-mediated lipoinitiation limits the bioengineering of NRPSs to obtain novel derivatives with desired acyl chains. Here, we show that the acyl chains of the lipopeptides rhizomide, holrhizin, and glidobactin were modified by engineering the starter condensation domain, suggesting a workable approach to change the acyl chain. Based on the structure of the mutated starter condensation domain of rhizomide biosynthetic enzyme RzmA in complex with octanoyl-CoA and related point mutation experiments, we identify a set of residues responsible for the selectivity of substrate acyl chains and extend the acyl chains from acetyl to palmitoyl. Furthermore, we illustrate three possible conformational states of starter condensation domains during the reaction cycle of the lipoinitiation process. Our studies provide further insights into the mechanism of lipoinitiation and the engineering of nonribosomal peptide synthetases.
Topics: Acylation; Amino Acid Sequence; Lipids; Lipopeptides; Models, Molecular; Peptide Biosynthesis, Nucleic Acid-Independent; Point Mutation; Protein Domains; Protein Engineering; Substrate Specificity
PubMed: 33436600
DOI: 10.1038/s41467-020-20548-8 -
Nucleic Acids Research May 2021Ribosomes are evolutionary conserved ribonucleoprotein complexes that function as two separate subunits in all kingdoms. During translation initiation, the two subunits...
Ribosomes are evolutionary conserved ribonucleoprotein complexes that function as two separate subunits in all kingdoms. During translation initiation, the two subunits assemble to form the mature ribosome, which is responsible for translating the messenger RNA. When the ribosome reaches a stop codon, release factors promote translation termination and peptide release, and recycling factors then dissociate the two subunits, ready for use in a new round of translation. A tethered ribosome, called Ribo-T, in which the two subunits are covalently linked to form a single entity, was recently described in Escherichia coli. A hybrid ribosomal RNA (rRNA) consisting of both the small and large subunit rRNA sequences was engineered. The ribosome with inseparable subunits generated in this way was shown to be functional and to sustain cell growth. Here, we investigated the translational properties of Ribo-T. We analyzed its behavior during amino acid misincorporation, -1 or +1 frameshifting, stop codon readthrough, and internal translation initiation. Our data indicate that covalent attachment of the two subunits modifies the properties of the ribosome, altering its ability to initiate and terminate translation correctly.
Topics: Codon, Terminator; Frameshifting, Ribosomal; Peptide Chain Initiation, Translational; Peptide Chain Termination, Translational; Protein Biosynthesis; RNA, Transfer; Ribosomes
PubMed: 33950196
DOI: 10.1093/nar/gkab259 -
European Journal of Biochemistry Aug 1990Peptide antibiotics are known to contain non-protein amino acids, D-amino acids, hydroxy acids, and other unusual constituents. In addition they may be modified by... (Review)
Review
Peptide antibiotics are known to contain non-protein amino acids, D-amino acids, hydroxy acids, and other unusual constituents. In addition they may be modified by N-methylation and cyclization reactions. Their biosynthetic origin has been connected in many cases to an enzymatic system referred to as the 'thiotemplate multienzymic mechanism'. This mechanism includes the activation of the constituent residues as adenylates on the enzymic template, the acylation of specific template thiol groups, epimerization or N-methylation at this thioester stage, and polymerization in the sequence directed by the multienzymic structure with the aid of 4'-phosphopantetheine as a cofactor, including possible cyclization or terminal modification reactions. The reaction sequences leading to gramicidin S, tyrocidine, cyclosporine, bacitracin, polymyxin, actinomycin, enniatin, beauvericin, delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine and linear gramicidin are discussed. The structures of the multienzymes, their genetic organization, the biological functions of these peptides and results on related systems are discussed.
Topics: Amino Acid Sequence; Anti-Bacterial Agents; Molecular Sequence Data; Peptide Biosynthesis; Ribosomes
PubMed: 2205497
DOI: 10.1111/j.1432-1033.1990.tb19188.x