Formylglycine, a Post-Translationally Generated Residue with Unique Catalytic Capabilities and Biotechnology Applications

Appel, Mason J.; Bertozzi, Carolyn R.

doi:10.1021/cb500897w

Cited by 153 publications

(147 citation statements)

References 106 publications

(272 reference statements)

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“…Crystallographic structure of FGE (Bond et al, 1997;von Bülow et al, 2001) and anSME (Goldman et al, 2013) demonstrate that their mechanisms differ in the use or non-use of oxygen molecules, respectively. A recent review covers the state of knowledge on these enzymes as well as their potential biotechnological applications (Appel and Bertozzi, 2015).…”

Section: Mechanism and Structure Of Sulfatasesmentioning

confidence: 99%

Marine Polysaccharide Sulfatases

Helbert

2017

Front. Mar. Sci.

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Polysaccharides are the most abundant and the most complex organic molecules in the ocean. In contrast to land polysaccharides, many marine polysaccharides are highly sulfated; in particular, the cell walls of macroalgae harbor a high diversity of sulfated polysaccharides (carrageenans, agarans, fucoidans, ulvans, etc.). These sulfated polysaccharides, biosynthesized by macroalgal primary producers, represent an important food source for heterotrophic organisms. Their biodegradation requires a set of enzymes that can cleave the glycosidic linkages of the carbohydrate backbone (called glycoside hydrolases) and the sulfate ester groups (called polysaccharide sulfatases). This review first provides on overview of the current state of knowledge on the classification and mechanisms of sulfatases in general. Then, based on an exploration of marine genomic and metagenomics data that reveals the diversity of carbohydrate sulfatases, it focuses on strategies to predict these sulfatases. In particular, the modularity of sulfatases and their location in marine polysaccharide utilization loci (PUL) provide clues as to their potential substrates and can drive future functional assays. Finally, the review underscores the low number of currently biochemically characterized marine carbohydrate sulfatases (e.g., agarases, carrageenanases, and fucose sulfatases). Bottlenecks encountered in studies on sulfatases likely lie in the difficulties in purifying them and producing them in heterologous systems.

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Section: Mechanism and Structure Of Sulfatasesmentioning

confidence: 99%

Marine Polysaccharide Sulfatases

Helbert

2017

Front. Mar. Sci.

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“…The nucleophilic cysteine of papain was chemically mutated via the intermediate formylglycine. Notably, formylglycine has itself since been recognized as a desirable residue, both as a natural PTM (derived enzymatically from Cys or Ser) mediating sulfate ester hydrolysis at the active site of type I sulfatases, and as a uniquely reactive aldehyde tag for further modification 11. Synthetically, chemical mutation of papain by Clark and Lowe commenced with selective alkylation of the reactive active‐site Cys using phenacyl bromide.…”

Section: The Early Years: βγ‐Carbon–heteroatom Bond Formationmentioning

confidence: 99%

From Chemical Mutagenesis to Post‐Expression Mutagenesis: A 50 Year Odyssey

2016

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Site‐directed (gene) mutagenesis has been the most useful method available for the conversion of one amino acid residue of a given protein into another. Until relatively recently, this strategy was limited to the twenty standard amino acids. The ongoing maturation of stop codon suppression and related technologies for unnatural amino acid incorporation has greatly expanded access to nonstandard amino acids by expanding the scope of the translational apparatus. However, the necessity for translation of genetic changes restricts the diversity of residues that may be incorporated. Herein we highlight an alternative approach, termed post‐expression mutagenesis, which operates at the level of the very functional biomolecules themselves. Using the lens of retrosynthesis, we highlight prospects for new strategies in protein modification, alteration, and construction which will enable protein science to move beyond the constraints of the “translational filter” and lead to a true synthetic biology.

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“…The Mycobacterium tuberculosis formyl-glycine generating enzyme (FGE) specifically catalyzes the co-translational oxidation of the cysteine residue on the hexapeptide sequence LCTPSR to an aldehyde containing a Cα-formylglycine (fGly) residue, which can be then labeled in vitro (Appel and Bertozzi, 2015, Liu et al, 2015, Rabuka et al, 2012, Shi et al, 2012, Ghoneim and Spies, 2014). The FGE recognition sequence can be genetically encoded into a protein, which is then co-expressed with FGE, both in eukaryotic as well as prokaryotic expression systems (Rabuka et al, 2012).…”

Section: Observing Macromolecular Interactions Using Single-molecumentioning

confidence: 99%

Quantifying the Assembly of Multicomponent Molecular Machines by Single-Molecule Total Internal Reflection Fluorescence Microscopy

Boehm

Subramanyam

Ghoneim

et al. 2016

Methods in Enzymology

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Large, dynamic macromolecular complexes play essential roles in many cellular processes. Knowing how the components of these complexes associate with one another and undergo structural rearrangements is critical to understanding how they function. Single-molecule total internal reflection fluorescence (TIRF) microscopy is a powerful approach for addressing these fundamental issues. In this article, we first discuss single-molecule TIRF microscopes and strategies to immobilize and fluorescently label macromolecules. We then review the use of single-molecule TIRF microscopy to study the formation of binary macromolecular complexes using one-color imaging and inhibitors. We conclude with a discussion of the use of TIRF microscopy to examine the formation of higher-order (i.e., ternary, quaternary, etc.) complexes using multi-color setups. The focus throughout this article is on experimental design, controls, data acquisition, and data analysis. We hope that single-molecule TIRF microscopy, which has largely been the province of specialists, will soon become as common in the tool box of biophysicists and biochemists as structural approaches has become today.

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Formylglycine, a Post-Translationally Generated Residue with Unique Catalytic Capabilities and Biotechnology Applications

Cited by 153 publications

References 106 publications

Marine Polysaccharide Sulfatases

Marine Polysaccharide Sulfatases

From Chemical Mutagenesis to Post‐Expression Mutagenesis: A 50 Year Odyssey

Quantifying the Assembly of Multicomponent Molecular Machines by Single-Molecule Total Internal Reflection Fluorescence Microscopy

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