2.0Å Resolution Crystal Structures of the Ternary Complexes of Human Phenylalanine Hydroxylase Catalytic Domain with Tetrahydrobiopterin and 3-(2-Thienyl)-l-alanine or l-Norleucine: Substrate Specificity and Molecular Motions Related to Substrate Binding

2003 ◽  
Vol 333 (4) ◽  
pp. 747-757 ◽  
Author(s):  
Ole Andreas Andersen ◽  
Anne J Stokka ◽  
Torgeir Flatmark ◽  
Edward Hough
Keyword(s):  
Substrate Specificity ◽  
Crystal Structures ◽  
Catalytic Domain ◽  
Phenylalanine Hydroxylase ◽  
Substrate Binding ◽  
Ternary Complexes ◽  
Molecular Motions ◽  
Human Phenylalanine Hydroxylase

Crystal structure of the catalytic domain of human phenylalanine hydroxylase reveals the structural basis for phenylketonuria

1997 ◽  
Vol 4 (12) ◽  
pp. 995-1000 ◽  
Author(s):  
Heidi Erlandsen ◽  
Fabrizia Fusetti ◽  
Aurora Martinez ◽  
Edward Hough ◽  
Torgeir Flatmark ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Catalytic Domain ◽  
Phenylalanine Hydroxylase ◽  
Structural Basis ◽  
Human Phenylalanine Hydroxylase

scholarly journals Crystal Structures of Yeast β-Alanine Synthase Complexes Reveal the Mode of Substrate Binding and Large Scale Domain Closure Movements

2007 ◽  
Vol 282 (49) ◽  
pp. 36037-36047 ◽  
Author(s):  
Stina Lundgren ◽  
Birgit Andersen ◽  
Jure Piškur ◽  
Doreen Dobritzsch
Keyword(s):  
Crystal Structures ◽  
Large Scale ◽  
Catalytic Domain ◽  
Substrate Binding ◽  
Salt Bridge ◽  
Site Directed Mutagenesis ◽  
Wild Type ◽  
Dimerization Domain ◽  
Binding Residues ◽  
Present Site

β-Alanine synthase is the final enzyme of the reductive pyrimidine catabolic pathway, which is responsible for the breakdown of uracil and thymine in higher organisms. The fold of the homodimeric enzyme from the yeast Saccharomyces kluyveri identifies it as a member of the AcyI/M20 family of metallopeptidases. Its subunit consists of a catalytic domain harboring a di-zinc center and a smaller dimerization domain. The present site-directed mutagenesis studies identify Glu159 and Arg322 as crucial for catalysis and His262 and His397 as functionally important but not essential. We determined the crystal structures of wild-type β-alanine synthase in complex with the reaction product β-alanine, and of the mutant E159A with the substrate N-carbamyl-β-alanine, revealing the closed state of a dimeric AcyI/M20 metallopeptidase-like enzyme. Subunit closure is achieved by a ∼30° rigid body domain rotation, which completes the active site by integration of substrate binding residues that belong to the dimerization domain of the same or the partner subunit. Substrate binding is achieved via a salt bridge, a number of hydrogen bonds, and coordination to one of the zinc ions of the di-metal center.

scholarly journals Insights into Substrate Specificity of NlpC/P60 Cell Wall Hydrolases Containing Bacterial SH3 Domains

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Qingping Xu ◽  
Dominique Mengin-Lecreulx ◽  
Xueqian W. Liu ◽  
Delphine Patin ◽  
Carol L. Farr ◽  
...  
Keyword(s):  
Cell Wall ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Catalytic Domain ◽  
Substrate Binding ◽  
Diaminopimelic Acid ◽  
Single Mutation ◽  
Molecular Architecture ◽  
Substrate Specificities ◽  
Cell Wall Hydrolases

ABSTRACTBacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. These enzymes all have γ-d-Glu-A2pm (A2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structure consisting of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation.IMPORTANCEPeptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural analysis of three modular NlpC/P60 hydrolases, one lysin, and two recycling enzymes, show that they may have evolved from a common molecular architecture, where the substrate preference is modulated by local changes. These results also suggest that new pathways for recycling PG turnover products, such as tracheal cytotoxin, may have evolved in bacteria in the human gut microbiome that involve NlpC/P60 cell wall hydrolases.

Structural characterization of a mitochondrial 3-ketoacyl-CoA (T1)-like thiolase fromMycobacterium smegmatis

2015 ◽  
Vol 71 (12) ◽  
pp. 2479-2493 ◽  
Author(s):  
Neelanjana Janardan ◽  
Rajesh K. Harijan ◽  
Tiila-Riikka Kiema ◽  
Rikkert K. Wierenga ◽  
M. R. N. Murthy
Keyword(s):  
Substrate Specificity ◽  
Crystal Structures ◽  
Structural Characterization ◽  
Mycobacterium Smegmatis ◽  
Substrate Binding ◽  
Residual Activity ◽  
Crystal Forms ◽  
Tetramerization Domain ◽  
Acyl Chains

Thiolases catalyze the degradation and synthesis of 3-ketoacyl-CoA molecules. Here, the crystal structures of a T1-like thiolase (MSM-13 thiolase) fromMycobacterium smegmatisin apo and liganded forms are described. Systematic comparisons of six crystallographically independent unliganded MSM-13 thiolase tetramers (dimers of tight dimers) from three different crystal forms revealed that the two tight dimers are connected to a rigid tetramerization domainviaflexible hinge regions, generating an asymmetric tetramer. In the liganded structure, CoA is bound to those subunits that are rotated towards the tip of the tetramerization loop of the opposing dimer, suggesting that this loop is important for substrate binding. The hinge regions responsible for this rotation occur near Val123 and Arg149. The Lα1–covering loop–Lα2 region, together with the Nβ2–Nα2 loop of the adjacent subunit, defines a specificity pocket that is larger and more polar than those of other tetrameric thiolases, suggesting that MSM-13 thiolase has a distinct substrate specificity. Consistent with this finding, only residual activity was detected with acetoacetyl-CoA as the substrate in the degradative direction. No activity was observed with acetyl-CoA in the synthetic direction. Structural comparisons with other well characterized thiolases suggest that MSM-13 thiolase is probably a degradative thiolase that is specific for 3-ketoacyl-CoA molecules with polar, bulky acyl chains.

scholarly journals Structural characteristics of an insect group I chitinase, an enzyme indispensable to moulting

2014 ◽  
Vol 70 (4) ◽  
pp. 932-942 ◽  
Author(s):  
Lei Chen ◽  
Tian Liu ◽  
Yong Zhou ◽  
Qi Chen ◽  
Xu Shen ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Catalytic Domain ◽  
Structural Characteristics ◽  
Substrate Binding ◽  
Hydrolysis Product ◽  
Binding Activity ◽  
Boat Conformation ◽  
Group I ◽  
Binding Cleft ◽  
Insect Group

Insects possess a greater number of chitinases than any other organisms. This work is the first report of unliganded and oligosaccharide-complexed crystal structures of the insect chitinaseOfChtI fromOstrinia furnacalis, which is essential to moulting. The obtained crystal structures were solved at resolutions between 1.7 and 2.2 Å. A structural comparison with other chitinases revealed thatOfChtI contains a long substrate-binding cleft similar to the bacterial chitinaseSmChiB fromSerratia marcescens. However, unlike the exo-actingSmChiB, which has a blocked and tunnel-like cleft,OfChtI possesses an open and groove-like cleft. The complexed structure of the catalytic domain ofOfChtI (OfChtI-CAD) with (GlcNAc)2/3indicates that the reducing sugar at subsite −1 is in an energetically unfavoured `boat' conformation, a state that possibly exists just before the completion of catalysis. BecauseOfChtI is known to act from nonreducing ends, (GlcNAc)3would be a hydrolysis product of (GlcNAc)6, suggesting thatOfChtI possesses an endo enzymatic activity. Furthermore, a hydrophobic plane composed of four surface-exposed aromatic residues is adjacent to the entrance to the substrate-binding cleft. Mutations of these residues greatly impair the chitin-binding activity, indicating that this hydrophobic plane endowsOfChtI-CAD with the ability to anchor chitin. This work reveals the unique structural characteristics of an insect chitinase.

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