Mechanistic Investigations on Microbial Type I Terpene Synthases through Site-directed Mutagenesis

Synthesis ◽  
2021 ◽  
Author(s):  
Houchao Xu ◽  
Jeroen Dickschat
Keyword(s):  
Crystal Structures ◽  
Site Directed Mutagenesis ◽  
Terpene Synthase ◽  
Directed Mutagenesis ◽  
Type I ◽  
Terpene Synthases ◽  
Conserved Motifs ◽  
Sequence Identity ◽  
The Past ◽  
Mechanistic Investigations

During the past three decades many terpene synthases have been characterised from all kingdoms of life. The type I of these enzymes from bacteria, fungi and protists commonly exhibit several highly conserved motifs and single residues, and the available crystal structures show a shared -helical fold, while the overall sequence identity is generally low. Several enzymes have been studied by site-directed mutagenesis, giving valuable insights into terpene synthase catalysis and the intriguing mechanisms of terpene synthases. Some mutants are also preparatively useful and give higher yields than the wildtype or a different product that is otherwise difficult to access. The accumulated knowledge obtained from these studies is presented and discussed in this review.

β-Amyrin biosynthesis: catalytic mechanism and substrate recognition

2017 ◽  
Vol 15 (14) ◽  
pp. 2869-2891 ◽  
Author(s):  
Tsutomu Hoshino
Keyword(s):  
Catalytic Mechanism ◽  
Substrate Recognition ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
The Past ◽  
Substrate Analog ◽  
Analog Experiments

In the past five years, there have been remarkable advances in the study of β-amyrin synthase. This review outlines the catalytic mechanism and substrate recognition in β-amyrin biosynthesis, which have been attained by the site-directed mutagenesis and substrate analog experiments.

scholarly journals Domain sliding of two Staphylococcus aureus N-acetylglucosaminidases enables their substrate-binding prior to its catalysis

2020 ◽  
Vol 3 (1) ◽  
Author(s):  
Sara Pintar ◽  
Jure Borišek ◽  
Aleksandra Usenik ◽  
Andrej Perdih ◽  
Dušan Turk
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
Drug Targets ◽  
Substrate Binding ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Human Pathogen ◽  
Active Site Cleft ◽  
Novel Drug ◽  
Novel Drug Targets

AbstractTo achieve productive binding, enzymes and substrates must align their geometries to complement each other along an entire substrate binding site, which may require enzyme flexibility. In pursuit of novel drug targets for the human pathogen S. aureus, we studied peptidoglycan N-acetylglucosaminidases, whose structures are composed of two domains forming a V-shaped active site cleft. Combined insights from crystal structures supported by site-directed mutagenesis, modeling, and molecular dynamics enabled us to elucidate the substrate binding mechanism of SagB and AtlA-gl. This mechanism requires domain sliding from the open form observed in their crystal structures, leading to polysaccharide substrate binding in the closed form, which can enzymatically process the bound substrate. We suggest that these two hydrolases must exhibit unusual extents of flexibility to cleave the rigid structure of a bacterial cell wall.

Structure and allosteric regulation of eukaryotic 6-phosphofructokinases

2013 ◽  
Vol 394 (8) ◽  
pp. 977-993 ◽  
Author(s):  
Torsten Schöneberg ◽  
Marco Kloos ◽  
Antje Brüser ◽  
Jürgen Kirchberger ◽  
Norbert Sträter
Keyword(s):  
Crystal Structures ◽  
Structural Information ◽  
Current Knowledge ◽  
Allosteric Regulation ◽  
Site Directed Mutagenesis ◽  
Structural Basis ◽  
Directed Mutagenesis ◽  
Special Focus ◽  
Molecular Architecture ◽  
Key Enzyme

Abstract Although the crystal structures of prokaryotic 6-phosphofructokinase, a key enzyme of glycolysis, have been available for almost 25 years now, structural information about the more complex and highly regulated eukaryotic enzymes is still lacking until now. This review provides an overview of the current knowledge of eukaryotic 6-phosphofructokinase based on recent crystal structures, kinetic analyses and site-directed mutagenesis data with special focus on the molecular architecture and the structural basis of allosteric regulation.

Comparative aspects of glycosyltransferases

2002 ◽  
Vol 69 ◽  
pp. 23-32 ◽  
Author(s):  
Christelle Breton ◽  
Helena Heissigerová ◽  
Charlotte Jeanneau ◽  
Jitka Moravcová ◽  
Anne Imberty
Keyword(s):  
Crystal Structures ◽  
Fold Recognition ◽  
The Other ◽  
Biological Functions ◽  
Direct Role ◽  
Conserved Motifs ◽  
Sequence Identity ◽  
Functional Aspects ◽  
The One ◽  
Limited Sequence

Glycosyltransferases, the enzymes that build oligosaccharides and glycoconjugates, have received much interest in recent years owing to their biological functions and their potential uses in biotechnology. Despite the fact that many glycosyltransferases recognize similar donor or acceptor substrates, there is surprisingly limited sequence identity between different classes. On the one hand, the glycosyltransferases are found in a large number of families, by sequence-based classification. On the other hand, only two structural folds have been identified among the fewer than one dozen glycosyltransferases that have been crystallized at present. Detection of conserved motifs that have a direct role in the functional aspects of glycosyltransferases is one approach for identifying remote similarity. With the availability of more crystal structures, the use of the fold-recognition approach is also very promising.

1, 10/1, 11 Cyclization catalyzed by diverged plant sesquiterpene synthases is dependent on a single residue

2021 ◽  
Author(s):  
Xin Fang ◽  
Jin-Quan Huang ◽  
Dong-Mei Li ◽  
Jian-Xu Li ◽  
Jia-Ling Lin ◽  
...  
Keyword(s):  
Phylogenetic Analyses ◽  
Site Directed Mutagenesis ◽  
Terpene Synthase ◽  
Directed Mutagenesis

The exquisite chemodiversity of terpenoids are the product of the large diverse terpene synthase (TPS) superfamily. Here, by using structural and phylogenetic analyses and site-directed mutagenesis, we identified a residue...

Crystal structures and functional studies clarify substrate selectivity and catalytic residues for the unique orphan enzyme N-acetyl-D-mannosamine dehydrogenase

2014 ◽  
Vol 462 (3) ◽  
pp. 499-511 ◽  
Author(s):  
Agustín Sola-Carvajal ◽  
Fernando Gil-Ortiz ◽  
Francisco García-Carmona ◽  
Vicente Rubio ◽  
Álvaro Sánchez-Ferrer
Keyword(s):  
Substrate Specificity ◽  
Crystal Structures ◽  
Functional Site ◽  
Site Directed Mutagenesis ◽  
Short Chain ◽  
Directed Mutagenesis ◽  
Substrate Selectivity ◽  
Functional Studies ◽  
Short Chain Dehydrogenase ◽  
Orphan Enzyme

Functional site-directed mutagenesis and crystallographic studies with N-acetyl-D-mannosamine dehydrogenase reveal a homotetramer, a short-chain dehydrogenase/reductase subunit fold and a highly developed C-terminal tail interlinking subunit. The structures of the complexes explain substrate specificity. The four residues forming the catalytic tetrad are identified.

scholarly journals Site-Directed Mutagenesis Analysis of Amino Acids Critical for Activity of the Type I Signal Peptidase of the Archaeon Methanococcus voltae

2005 ◽  
Vol 187 (3) ◽  
pp. 1188-1191 ◽  
Author(s):  
Sonia L. Bardy ◽  
Sandy Y. M. Ng ◽  
David S. Carnegie ◽  
Ken F. Jarrell
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Aspartic Acid ◽  
Site Directed Mutagenesis ◽  
Signal Peptidase ◽  
Directed Mutagenesis ◽  
Type I ◽  
Aspartic Acid Residue ◽  
Conserved Residues ◽  
Methanococcus Voltae

ABSTRACT Site-directed mutagenesis studies of the signal peptidase of the methanogenic archaeon Methanococcus voltae identified three conserved residues (Ser52, His122, and Asp148) critical for activity. The requirement for one conserved aspartic acid residue distinguishes the archaeal enzyme from both the Escherichia coli and yeast Sec11 enzymes.

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