Evolution of Enzymatic Activities in the Orotidine 5‘-Monophosphate Decarboxylase Suprafamily:  Structural Basis for Catalytic Promiscuity in Wild-Type and Designed Mutants of 3-Keto-l-gulonate 6-Phosphate Decarboxylase†

Biochemistry ◽  
2005 ◽  
Vol 44 (6) ◽  
pp. 1816-1823 ◽  
Author(s):  
Eric L. Wise ◽  
Wen Shan Yew ◽  
Julie Akana ◽  
John A. Gerlt ◽  
Ivan Rayment
Keyword(s):  
Enzymatic Activities ◽  
Structural Basis ◽  
Catalytic Promiscuity ◽  
Wild Type ◽  
Monophosphate Decarboxylase

Discovery of natural product ovalicin sensitive type 1 methionine aminopeptidases: molecular and structural basis

2019 ◽  
Vol 476 (6) ◽  
pp. 991-1003 ◽  
Author(s):  
Vijaykumar Pillalamarri ◽  
Tarun Arya ◽  
Neshatul Haque ◽  
Sandeep Chowdary Bala ◽  
Anil Kumar Marapaka ◽  
...  
Keyword(s):  
Natural Product ◽  
Active Site ◽  
Covalent Modification ◽  
Structural Basis ◽  
Wild Type ◽  
Methionine Aminopeptidases ◽  
Synthetic Derivative ◽  
Dynamics Simulations

Abstract Natural product ovalicin and its synthetic derivative TNP-470 have been extensively studied for their antiangiogenic property, and the later reached phase 3 clinical trials. They covalently modify the conserved histidine in Type 2 methionine aminopeptidases (MetAPs) at nanomolar concentrations. Even though a similar mechanism is possible in Type 1 human MetAP, it is inhibited only at millimolar concentration. In this study, we have discovered two Type 1 wild-type MetAPs (Streptococcus pneumoniae and Enterococcus faecalis) that are inhibited at low micromolar to nanomolar concentrations and established the molecular mechanism. F309 in the active site of Type 1 human MetAP (HsMetAP1b) seems to be the key to the resistance, while newly identified ovalicin sensitive Type 1 MetAPs have a methionine or isoleucine at this position. Type 2 human MetAP (HsMetAP2) also has isoleucine (I338) in the analogous position. Ovalicin inhibited F309M and F309I mutants of human MetAP1b at low micromolar concentration. Molecular dynamics simulations suggest that ovalicin is not stably placed in the active site of wild-type MetAP1b before the covalent modification. In the case of F309M mutant and human Type 2 MetAP, molecule spends more time in the active site providing time for covalent modification.

Regulation of myosin 5a and myosin 7a

2011 ◽  
Vol 39 (5) ◽  
pp. 1136-1141 ◽  
Author(s):  
Verl B. Siththanandan ◽  
James R. Sellers
Keyword(s):  
Cellular Function ◽  
Enzymatic Activities ◽  
Motor Domain ◽  
Present Review ◽  
Structural Basis ◽  
Ion Binding ◽  
Inactive State ◽  
Kinetic Mechanisms ◽  
Myosin Superfamily

The myosin superfamily is diverse in its structure, kinetic mechanisms and cellular function. The enzymatic activities of most myosins are regulated by some means such as Ca2+ ion binding, phosphorylation or binding of other proteins. In the present review, we discuss the structural basis for the regulation of mammalian myosin 5a and Drosophila myosin 7a. We show that, although both myosins have a folded inactive state in which domains in the myosin tail interact with the motor domain, the details of the regulation of these two myosins differ greatly.

scholarly journals Catalytic promiscuity and the evolution of new enzymatic activities

1999 ◽  
Vol 6 (4) ◽  
pp. R91-R105 ◽  
Author(s):  
Patrick J O'Brien ◽  
Daniel Herschlag
Keyword(s):  
Enzymatic Activities ◽  
Catalytic Promiscuity

scholarly journals Structural Basis and Functional Consequence of Helicobacter pylori CagA Multimerization in Cells

2006 ◽  
Vol 281 (43) ◽  
pp. 32344-32352 ◽  
Author(s):  
Shumei Ren ◽  
Hideaki Higashi ◽  
Huaisheng Lu ◽  
Takeshi Azuma ◽  
Masanori Hatakeyama
Keyword(s):  
Helicobacter Pylori ◽  
Tyrosine Phosphorylation ◽  
Cell Shape ◽  
Host Cells ◽  
Structural Basis ◽  
Sequence Motif ◽  
Wild Type ◽  
Src Homology 2 ◽  
Src Homology ◽  
In Cells

Helicobacter pylori cagA-positive strains are associated with gastric adenocarcinoma. The cagA gene product CagA is delivered into gastric epithelial cells where it localizes to the plasma membrane and undergoes tyrosine phosphorylation at the EPIYA-repeat region, which contains the EPIYA-A segment, EPIYA-B segment, and Western CagA-specific EPIYA-C or East Asian CagA-specific EPIYA-D segment. In host cells, CagA specifically binds to and deregulates SHP-2 phosphatase via the tyrosine-phosphorylated EPIYA-C or EPIYA-D segment, thereby inducing an elongated cell shape known as the hummingbird phenotype. In this study, we found that CagA multimerizes in cells in a manner independent of its tyrosine phosphorylation. Using a series of CagA mutants, we identified a conserved amino acid sequence motif (FPLXRXXXVXDLSKVG), which mediates CagA multimerization, within the EPIYA-C segment as well as in a sequence that located immediately downstream of the EPIYA-C or EPIYA-D segment. We also found that a phosphorylation-resistant CagA, which multimerizes but cannot bind SHP-2, inhibits the wild-type CagA-SHP-2 complex formation and abolishes induction of the hummingbird phenotype. Thus, SHP-2 binds to a preformed and tyrosinephosphorylated CagA multimer via its two Src homology 2 domains. These results, in turn, indicate that CagA multimerization is a prerequisite for CagA-SHP-2 interaction and subsequent deregulation of SHP-2. The present work raises the possibility that inhibition of CagA multimerization abolishes pathophysiological activities of CagA that promote gastric carcinogenesis.

Structural basis of the transactivation deficiency of the human PPARγ F360L mutant associated with familial partial lipodystrophy

2014 ◽  
Vol 70 (7) ◽  
pp. 1965-1976 ◽  
Author(s):  
Clorinda Lori ◽  
Alessandra Pasquo ◽  
Roberta Montanari ◽  
Davide Capelli ◽  
Valerio Consalvi ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Conformational Changes ◽  
Partial Agonist ◽  
Salt Bridge ◽  
Van Der Waals Interactions ◽  
Structural Basis ◽  
Wild Type ◽  
Partial Lipodystrophy ◽  
X Ray ◽  
Familial Partial Lipodystrophy

The peroxisome proliferator-activated receptors (PPARs) are transcription factors that regulate glucose and lipid metabolism. The role of PPARs in several chronic diseases such as type 2 diabetes, obesity and atherosclerosis is well known and, for this reason, they are the targets of antidiabetic and hypolipidaemic drugs. In the last decade, some rare mutations in human PPARγ that might be associated with partial lipodystrophy, dyslipidaemia, insulin resistance and colon cancer have emerged. In particular, the F360L mutant of PPARγ (PPARγ2 residue 388), which is associated with familial partial lipodystrophy, significantly decreases basal transcriptional activity and impairs stimulation by synthetic ligands. To date, the structural reason for this defective behaviour is unclear. Therefore, the crystal structure of PPARγ F360L together with the partial agonist LT175 has been solved and the mutant has been characterized by circular-dichroism spectroscopy (CD) in order to compare its thermal stability with that of the wild-type receptor. The X-ray analysis showed that the mutation induces dramatic conformational changes in the C-terminal part of the receptor ligand-binding domain (LBD) owing to the loss of van der Waals interactions made by the Phe360 residue in the wild type and an important salt bridge made by Arg357, with consequent rearrangement of loop 11/12 and the activation function helix 12 (H12). The increased mobility of H12 makes the binding of co-activators in the hydrophobic cleft less efficient, thereby markedly lowering the transactivation activity. The spectroscopic analysis in solution and molecular-dynamics (MD) simulations provided results which were in agreement and consistent with the mutant conformational changes observed by X-ray analysis. Moreover, to evaluate the importance of the salt bridge made by Arg357, the crystal structure of the PPARγ R357A mutant in complex with the agonist rosiglitazone has been solved.

scholarly journals Interactions of Pulmonary Collectins with Bordetella bronchiseptica and Bordetella pertussis Lipopolysaccharide Elucidate the Structural Basis of Their Antimicrobial Activities

2004 ◽  
Vol 72 (12) ◽  
pp. 7124-7130 ◽  
Author(s):  
Lyndsay M. Schaeffer ◽  
Francis X. McCormack ◽  
Huixing Wu ◽  
Alison A. Weiss
Keyword(s):  
Bordetella Pertussis ◽  
Lipid A ◽  
Antimicrobial Activities ◽  
Innate Immune ◽  
Bordetella Bronchiseptica ◽  
Structural Basis ◽  
Wild Type ◽  
Core Oligosaccharide ◽  
The Core ◽  
O Antigen

ABSTRACT Surfactant proteins A (SP-A) and D (SP-D) play an important role in the innate immune defenses of the respiratory tract. SP-A binds to the lipid A region of lipopolysaccharide (LPS), and SP-D binds to the core oligosaccharide region. Both proteins induce aggregation, act as opsonins for neutrophils and macrophages, and have direct antimicrobial activity. Bordetella pertussis LPS has a branched core structure and a nonrepeating terminal trisaccharide. Bordetella bronchiseptica LPS has the same structure, but lipid A is palmitoylated and there is a repeating O-antigen polysaccharide. The ability of SP-A and SP-D to agglutinate and permeabilize wild-type and LPS mutants of B. pertussis and B. bronchiseptica was examined. Previously, wild-type B. pertussis was shown to resist the effects of SP-A; however, LPS mutants lacking the terminal trisaccharide were susceptible to SP-A. In this study, SP-A was found to aggregate and permeabilize a B. bronchiseptica mutant lacking the terminal trisaccharide, while wild-type B. bronchiseptica and mutants lacking only the palmitoyl transferase or O antigen were resistant to SP-A. Wild-type B. pertussis and B. bronchiseptica were both resistant to SP-D; however, LPS mutants of either strain lacking the terminal trisaccharide were aggregated and permeabilized by SP-D. We conclude that the terminal trisaccharide protects Bordetella species from the bactericidal functions of SP-A and SP-D. The O antigen and palmitoylated lipid A of B. bronchiseptica play no role in this resistance.

Structural Basis of the Suppressed Catalytic Activity of Wild-type Human Glutathione Transferase T1-1 Compared to its W234R Mutant

2006 ◽  
Vol 355 (1) ◽  
pp. 96-105 ◽  
Author(s):  
Kaspars Tars ◽  
Anna-Karin Larsson ◽  
Abeer Shokeer ◽  
Birgit Olin ◽  
Bengt Mannervik ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Glutathione Transferase ◽  
Structural Basis ◽  
Wild Type

scholarly journals Substrate distortion in the catalysis of orotidine monophosphate decarboxylase

2014 ◽  
Vol 70 (a1) ◽  
pp. C447-C447
Author(s):  
Masahiro Fujihashi ◽  
Toyokazu Ishida ◽  
Shingo Kuroda ◽  
Kazuya Mito ◽  
Lakshmi Kotra ◽  
...  
Keyword(s):  
Computational Simulation ◽  
Complex Surface ◽  
Pyrimidine Ring ◽  
Wild Type ◽  
Wild Type Enzyme ◽  
Surface Plasmon Resonance Analysis ◽  
Resonance Analysis ◽  
Transition State Stabilization ◽  
Monophosphate Decarboxylase ◽  
Orotidine Monophosphate Decarboxylase

One way for enzymes to affect reactions they catalyze is through transition state stabilization. Another factor to be considered is the contribution of substrate distortion, although it has been thoroughly described for only a few enzymes. We have a longstanding interest in the reaction mechanism of orotidine monophosphate decarboxylase (ODCase) and determined various crystal structures bound with distorted substrates at around 1.5 Å resolution. The enzyme is known as one of the most proficient enzymes, which accelerates the decarboxylation of orotidine 5'-monophosphate (OMP) to form uridine 5'-monophosphate (UMP) by 17 orders of magnitude. One argument against the contribution of substrate distortion to the ODCase reaction is the weak affinity of UMP. The distortions observed so far all appear at the C6-substituent of the pyrimidine ring, which corresponds to the carboxylate of OMP. Since the carboxylate is removed by the reaction, the product UMP should bind more tightly to ODCase than OMP, if the distortion of C6-substituent contributes to the catalysis. In order to investigate this inconsistency, we determined the crystal structure of ODCase with UMP at atomic resolution (1.03 Å). The structure showed an unfavorable interaction between UMP and the catalytic residue K72, an interaction considered to be absent in the OMP complex. Surface plasmon resonance analysis indicated that UMP binds stronger to the K72A mutant than to the wild-type enzyme by 5 orders of magnitude. These analyses invalidate the argument against a contribution of substrate distortion to ODCase catalysis. Finally, we estimated how much the distortion contributes to the catalysis using computational simulation methods. The results indicated that 10-15% decrease of the ΔΔG‡ value is contributed by substrate distortion.

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