Open Access. Powered by Scholars. Published by Universities.®
Physical Sciences and Mathematics Commons™
Open Access. Powered by Scholars. Published by Universities.®
- Keyword
-
- Cobalt (6)
- Zinc (6)
- Aminopeptidase (4)
- Hydrolysis (4)
- Kinetics (4)
-
- Electron paramagnetic resonance (3)
- Electronic absorption (3)
- Aeromonas proteolytica (2)
- EPR (2)
- Enzyme inhibition (2)
- Manganese (2)
- Methionine aminopeptidases (2)
- Peptide hydrolysis (2)
- Tris (2)
- 2 (1)
- 6-Diaminopimelic acid derivatives (1)
- Active site ligands (1)
- Aeromonas proteolytica (Vibrio proteolyticus) (1)
- Aminopeptidases (1)
- Antibiotics (1)
- Arrays (1)
- Atomic force microscopy (1)
- Bacterial adhesion (1)
- Bacterial binding (1)
- Bioinorganic Chemistry (1)
- Biomaterials (1)
- Biomedicine (1)
- Biosynthesis (1)
- Buffer inhibition (1)
- Capillary zone electrophoresis (1)
- File Type
Articles 31 - 60 of 81
Full-Text Articles in Physical Sciences and Mathematics
Exafs Studies Of Uteroferrin And Its Anion Complexes, Anne True, Robert Scarrow, Clayton Randall, Richard Holz, Lawrence Que
Exafs Studies Of Uteroferrin And Its Anion Complexes, Anne True, Robert Scarrow, Clayton Randall, Richard Holz, Lawrence Que
Richard C. Holz
Iron K-edge X-ray absorption data on the purple acid phosphatase from porcine uterus (uteroferrin, Uf) have been obtained for the native reduced enzyme and for the oxidized enzyme in its phosphate- and arsenate-bound forms. In all three complexes, the first sphere consists of 1.5 N/O at ∼1.94 Å, 4 N/O at ∼2.1 Å, and 0.5-1 N/O at ∼2.4 Å; in no complex is found an Fe-O bond of ∼1.8 Å which would derive from a μ-oxo bond. The ∼1.94-Å shell corresponds to Fe-OAr and Fe-μ-OH(or R) bonds. The ∼2.1-Å shell arises from histidine, carboxylate, oxoanion, and solvent ligation. The scatterer …
Mutation Of H63 And Its Catalytic Affect On The Methionine Aminopeptidase From Escherichia Coli, Sanghamitra Mitra, Brian Bennett, Richard C. Holz
Mutation Of H63 And Its Catalytic Affect On The Methionine Aminopeptidase From Escherichia Coli, Sanghamitra Mitra, Brian Bennett, Richard C. Holz
Richard C. Holz
In order to gain insight into the mechanistic role of a flexible exterior loop near the active site, made up of Y62, H63, G64, and Y65, that has been proposed to play an important role in substrate binding and recognition in the methionyl aminopeptidase from Escherichia coli (EcMetAP-I), the H63A enzyme was prepared. Mutation of H63 to alanine does not affect the ability of the enzyme to bind divalent metal ions. The specific activity of H63A EcMetAP-I was determined using four different substrates of varying lengths, namely, l-Met-p-NA, MAS, MGMM and MSSHRWDW. For the smallest/shortest …
Spectroscopic And Electrochemical Properties Of (Μ-Oxo)Diiron(Iii) Complexes Related To Diiron-Oxo Proteins. Structure Of [Fe2O(Tpa)2(Moo)4)](Clo4)2, Richard Holz, Timothy Elgren, Linda Pearce, Jian Zhang, Charles O'Connor, Lawrence Que
Spectroscopic And Electrochemical Properties Of (Μ-Oxo)Diiron(Iii) Complexes Related To Diiron-Oxo Proteins. Structure Of [Fe2O(Tpa)2(Moo)4)](Clo4)2, Richard Holz, Timothy Elgren, Linda Pearce, Jian Zhang, Charles O'Connor, Lawrence Que
Richard C. Holz
A series of (μ-oxo)diiron(III) complexes of tris(2-pyridylmethy1)amine (TPA), [Fe2O(TPA)2(L)] (C1O4)2, were synthesized and characterized where L represents the bridging tetraoxo anion ligands sulfate, phosphate, arsenate, vanadate, and molybdate. These tetraoxo anion complexes are the first (μ-oxo)diiron(III) complexes that reproduce the protein-tetraoxo anion stoichiometry found in purple acid phosphatases (PAPs). [Fe2O(TPA)2( MoO4)] (C104)2- CH3N (9) crystallizes in the monoclinic space group P21/n (a = 12.74(1) Å, b = 24.69(2) Å, c = 13.733(8) Å, β …
Laser-Induced Europium(Iii) Luminescence And Nmr Spectroscopic Characterization Of Macrocyclic Diaza Crown Ether Complexes Containing Carboxylate Ligating Groups, Richard Holz, Scott Klakamp, C. Chang, William Horrocks
Laser-Induced Europium(Iii) Luminescence And Nmr Spectroscopic Characterization Of Macrocyclic Diaza Crown Ether Complexes Containing Carboxylate Ligating Groups, Richard Holz, Scott Klakamp, C. Chang, William Horrocks
Richard C. Holz
The Eu3+ and Y3+ complexes of 1,10-diaza-4,7,13,16-tetraoxacyclooctadecane-N, N' - diacetic acid (K22DA), 1,7-diaza-4,10,13-trioxacyclopentadecane-N, N' - diacetic acid (K21DA), and the open-chain analogue ethylene glycol bis (ß-aminoethyl ether) - N,-N,N',N'-tetraacetic acid (EGTA) have been characterized in solution by using Eu3+ laser-induced luminescence and 1H and 13CNMR spectroscopy. All of these ligands form 1:1 complexes with Eu3+ in solution with the luminescence lifetimes in H2O and D2O providing the number of coordinated water molecules. Stepwise changes in the coordination environment of the Eu3+ and Y3+ ions were monitored spectroscopically for each complex as a function of temperature. In addition, the Eu3+ spectral …
Co-Catalytic Metallopeptidases As Pharmaceutical Targets, Richard Holz, Krzysztof Bzymek, Sabina Swierczek
Co-Catalytic Metallopeptidases As Pharmaceutical Targets, Richard Holz, Krzysztof Bzymek, Sabina Swierczek
Richard C. Holz
Understanding the reaction mechanism of co-catalytic metallopeptidases provides a starting point for the design and synthesis of new molecules that can be screened as potential pharmaceuticals. Many of the enzymes that contain co-catalytic metallo-active sites play important roles in cellular processes such as tissue repair, protein maturation, hormone level regulation, cell-cycle control and protein degradation. Therefore, these enzymes play central roles in several disease states including cancer, HIV, stroke, diabetes, bacterial infections, neurological processes, schizophrenia, seizure disorders, and amyotrophic lateral sclerosis. The mechanism of AAP, an aminopeptidase from Aeromonas proteolytica, is one of the best-characterized examples of a metallopeptidase containing …
Inhibition Of The Aminopeptidase From Aeromonas Proteolytica By L-Leucinephosphonic Acid, A Transition State Analogue Of Peptide Hydrolysis, Brian Bennett, Richard C. Holz
Inhibition Of The Aminopeptidase From Aeromonas Proteolytica By L-Leucinephosphonic Acid, A Transition State Analogue Of Peptide Hydrolysis, Brian Bennett, Richard C. Holz
Richard C. Holz
No abstract provided.
Characterization Of The Structural And Electronic Properties Of Spin-Coupled Dinuclear Copper(Ii) Centers By Proton Nmr Spectroscopy, Julie Brink, Rachel Rose, Richard Holz
Characterization Of The Structural And Electronic Properties Of Spin-Coupled Dinuclear Copper(Ii) Centers By Proton Nmr Spectroscopy, Julie Brink, Rachel Rose, Richard Holz
Richard C. Holz
The 1H NMR spectra of a series of well-characterized μ-phenoxo and μ-alkoxo spin-coupled dicopper(II) complexes have been investigated. The complexes studied were [Cu2(BPMP)(OH)]2+ (1) (BPMP = 2,6-bis[[bis(2-pyridylmethyl)amino]methyl]-4-methylphenol), [Cu2(CH3HXTA)(OH)]2- (2) (CH3HXTA = N,N ‘-(2-hydroxy-5-methyl-1,3-xylylene)bis(N-carboxymethylglycine), [Cu2(m-XYL)(OH)]2+ (3) (m-XYL = 2,6-bis[[bis(2-pyridylethyl)amino]methyl]phenol), and [Cu2(TBHP)(OAc)]2+ (4) (TBHP = N,N,N ‘,N ‘-tetrakis[(2-benzimidazolyl)methyl]-2-hydroxy-1,3-diaminopropane). The magnetic interactions of these complexes range from strongly antiferromagnetically to weakly ferromagnetically coupled. Both one- and two-dimensional (COSY) 1H NMR methods were used to facilitate the assignment of the hyperfine shifted 1H NMR signals of each complex. COSY experiments provide clear cross signals for resonances <200 Hz wide. These data have facilitated the …
The Catalytic Role Of Glutamate 151 In The Leucine Aminopeptidase From Aeromonas Proteolytica, Krzysztof Bzymek, Richard Holz
The Catalytic Role Of Glutamate 151 In The Leucine Aminopeptidase From Aeromonas Proteolytica, Krzysztof Bzymek, Richard Holz
Richard C. Holz
Glutamate 151 has been proposed to act as the general acid/base during the peptide hydrolysis reaction catalyzed by the co-catalytic metallohydrolase from Aeromonas proteolytica (AAP). However, to date, no direct evidence has been reported for the role of Glu-151 during catalytic turnover by AAP. In order to elucidate the catalytic role of Glu-151, altered AAP enzymes have been prepared in which Glu-151 has been substituted with a glutamine, an alanine, and an aspartate. The Michaelis constant (Km) does not change upon substitution to aspartate or glutamine, but the rate of the reaction changes drastically in the following order: glutamate (100% …
X-Ray Crystallographic Characterization Of The Co(Ii)-Substituted Tris-Bound Form Of The Aminopeptidase From Aeromonas Proteolytica, Petra Munih, Aaron Moulin, Carin Stamper, Brian Bennett, Dagmar Ringe, Gregory Petsko, Richard Holz
X-Ray Crystallographic Characterization Of The Co(Ii)-Substituted Tris-Bound Form Of The Aminopeptidase From Aeromonas Proteolytica, Petra Munih, Aaron Moulin, Carin Stamper, Brian Bennett, Dagmar Ringe, Gregory Petsko, Richard Holz
Richard C. Holz
The X-ray crystal structure of the Co(II)-loaded form of the aminopeptidase from Aeromonas proteolytica ([CoCo(AAP)]) was solved to 2.2 Å resolution. [CoCo(AAP)] folds into an α/β globular domain with a twisted β-sheet hydrophobic core sandwiched between α-helices, identical to [ZnZn(AAP)]. Co(II) binding to AAP does not introduce any major conformational changes to the overall protein structure and the amino acid residues ligated to the dicobalt(II) cluster in [CoCo(AAP)] are the same as those in the native Zn(II)-loaded structure with only minor perturbations in bond lengths. The Co(II)–Co(II) distance is 3.3 Å. Tris(hydroxymethyl)aminomethane (Tris) coordinates to the dinuclear Co(II) active site …
Kinetic And Spectroscopic Characterization Of The E134a- And E134d-Altered Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae, Ryan Davis, David Bienvenue, Sabina Swierczek, Danuta Gilner, Lakshman Rajagopal, Brian Bennett, Richard Holz
Kinetic And Spectroscopic Characterization Of The E134a- And E134d-Altered Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae, Ryan Davis, David Bienvenue, Sabina Swierczek, Danuta Gilner, Lakshman Rajagopal, Brian Bennett, Richard Holz
Richard C. Holz
Glutamate-134 (E134) is proposed to act as the general acid/base during the hydrolysis reaction catalyzed by the dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae. To date, no direct evidence has been reported for the role of E134 during catalytic turnover by DapE. In order to elucidate the catalytic role of E134, altered DapE enzymes were prepared in which E134 was substituted with an alanine and an aspartate residue. The Michaelis constant (K m) does not change upon substitution with aspartate but the rate of the reaction changes drastically in the following order: glutamate (100% activity), aspartate (0.09%), and alanine …
1-Butaneboronic Acid Binding To Aeromonas Proteolytica Aminopeptidase: A Case Of Arrested Development, Carin C. De Paola, Brian Bennett, Richard C. Holz, Dagmar Ringe, Gregory A. Petsko
1-Butaneboronic Acid Binding To Aeromonas Proteolytica Aminopeptidase: A Case Of Arrested Development, Carin C. De Paola, Brian Bennett, Richard C. Holz, Dagmar Ringe, Gregory A. Petsko
Richard C. Holz
Hydrolases containing two metal ions connected by a bridging ligand catalyze reactions important in carcinogensis, tissue repair, post-translational modification, control and regulation of biochemical pathways, and protein degradation. The aminopeptidase from Aeromonas proteolytica serves as a paradigm for the study of such bridged bimetallic proteases since its three-dimensional structure is known to very high resolution and its catalytic reaction is amenable to spectroscopic examination. Herein, we report the X-ray crystal structure at 1.9 Å resolution of AAP complexed with 1-butaneboronic acid (BuBA). This structure suggests that this complex represents a snapshot of the proteolytic reaction in an arrested form between …
Spectroscopic And Structural Characterization Of The Nine-Coordinate Adduct Of Tris(Dipivaloylmethanato)Europium(Iii) With 2,2':6',2"-Terpyridine, Richard Holz, L. Thompson
Spectroscopic And Structural Characterization Of The Nine-Coordinate Adduct Of Tris(Dipivaloylmethanato)Europium(Iii) With 2,2':6',2"-Terpyridine, Richard Holz, L. Thompson
Richard C. Holz
Tris(dipivaloylmethanato)(2,2':6',2"-terpyridine)europium(III), [Eu(DPM)3terpy], was synthesized from [EU(DPM)3] and 2,2':6',2"-terpyridine in carbon tetrachloride. The complex was characterized by infrared and luminescence spectroscopy. The infrared spectrum indicated all three nitrogen atoms of the terpyridine ligand were bound to the metal ion. The luminescence spectrum was recorded at 77 K and revealed a broad (15 cm-1) unresolvable single transition in the 5D0 -> 7F0 region. The 5D0 -> 7F1 region contained six bands and the 5D0 -> 7F2 region contained nine bands, indicating two distinct emitting metal centers. The luminescence spectrum suggested that the site symmetry of both metal centers is C1. The structure …
The Dimerization Domain In Dape Enzymes Is Required For Catalysis, Boguslaw Nocek, Anna Starus, Magdalena Makowska-Grzyska, Blanca Gutierrez, Stephen Sanchez, Robert Jedrzejczak, Jamey C. Mack, Kenneth W. Olsen, Andzrej Joachimiak, Richard C. Holz
The Dimerization Domain In Dape Enzymes Is Required For Catalysis, Boguslaw Nocek, Anna Starus, Magdalena Makowska-Grzyska, Blanca Gutierrez, Stephen Sanchez, Robert Jedrzejczak, Jamey C. Mack, Kenneth W. Olsen, Andzrej Joachimiak, Richard C. Holz
Richard C. Holz
The emergence of antibiotic-resistant bacterial strains underscores the importance of identifying new drug targets and developing new antimicrobial compounds. Lysine and meso-diaminopimelic acid are essential for protein production and bacterial peptidoglycan cell wall remodeling and are synthesized in bacteria by enzymes encoded within dap operon. Therefore dap enzymes may serve as excellent targets for developing a new class of antimicrobial agents. The dapE-encoded N-succinyl-L,L-diaminopimelic acid desuccinylase (DapE) converts N-succinyl-L,L-diaminopimelic acid to L,Ldiaminopimelic acid and succinate. The enzyme is composed of catalytic and dimerization domains, and belongs to the M20 peptidase family. To understand the specific role of each domain of …
Noesy Studies On The Fe(Iii)Co(Ii) Active Site Of The Purple Acid Phosphatase Uteroferrin, Richard Holz, Lawrence Que, Li-June Ming
Noesy Studies On The Fe(Iii)Co(Ii) Active Site Of The Purple Acid Phosphatase Uteroferrin, Richard Holz, Lawrence Que, Li-June Ming
Richard C. Holz
No abstract provided.
The 1.20 Å Resolution Crystal Structure Of The Aminopeptidase From Aeromonas Proteolytica Complexed With Tris: A Tale Of Buffer Inhibition, William Desmarais, David Bienvenue, Krzysztof Bzymek, Richard Holz, Gregory Petsko, Dagmar Ringe
The 1.20 Å Resolution Crystal Structure Of The Aminopeptidase From Aeromonas Proteolytica Complexed With Tris: A Tale Of Buffer Inhibition, William Desmarais, David Bienvenue, Krzysztof Bzymek, Richard Holz, Gregory Petsko, Dagmar Ringe
Richard C. Holz
The aminopeptidase from Aeromonas proteolytica (AAP) is a bridged bimetallic enzyme that removes the N-terminal amino acid from a peptide chain. To fully understand the metal roles in the reaction pathway of AAP we have solved the 1.20 Å resolution crystal structure of native AAP (PDB ID = 1LOK). The high-quality electron density maps showed a single Tris molecule chelated to the active site Zn2+, alternate side chain conformations for some side chains, a sodium ion that mediates a crystal contact, a surface thiocyanate ion, and several potential hydrogen atoms. In addition, the high precision of the atomic …
Spectroscopic Characterization Of A Series Of Europium(Iii) Amino Phosphonate Complexes In Solution, Richard Holz, Gretchen Meister, William Horrocks
Spectroscopic Characterization Of A Series Of Europium(Iii) Amino Phosphonate Complexes In Solution, Richard Holz, Gretchen Meister, William Horrocks
Richard C. Holz
The Eu3+ complexes of nitrilotris (methylenephosphonic acid) (ntp), ethylenediaminetetrakis (methylenephosphonic acid) (edtp), hexamethylenediaminetetrakis (methylenephosphonic acid) (hdtp), and diethylenetriaminepentakis (methylenephosphonic acid) (dtpp) have been characterized in solution by laser-induced Eu3+ luminescence spectroscopy. The latter three ligands form both 2:1 and 1:1 metal-ligand complexes, while ntp forms both 1:1 and 1:2 complexes at pH 6.0. The number of water molecules directly coordinating the metal ion was determined for each complex. The 7F0 -> 5D0 excitation spectra of edtp, hdtp, and dtpp reveal isomeric forms for both the 2:1 and 1:1 complexes while ntp exhibits only single species for both the 1:1 and …
Structure Of 2,9-Dimethyl-1,10-Phenanthroline Hemihydrate, Doyle Britton, Larry Thompson, Richard Holz
Structure Of 2,9-Dimethyl-1,10-Phenanthroline Hemihydrate, Doyle Britton, Larry Thompson, Richard Holz
Richard C. Holz
C14H12N2•½H2O, Mr = 217•27, tetragonal I41/a, a = 14•258 (3), c = 22•286 (4) Å, V = 4531 (3) Å3, Z = 16, Dx = 1•274 (1) g cm-3, Mo Kα radiation λ = 0•71073 Å, µ = 0•74 cm-1, F(000) = 1840, T = 297 K, R = 0•041 for 1196 unique observed reflections with I > 2σ(I). Pairs of dimethylphenanthroline molecules related by a twofold axis are bridged by water molecules lying on the twofold axis and H bonded to one of the N atoms in each molecule. The H bonds are long and far from linear: O—H 1•06 …
The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae Is A Dinuclear Metallohydrolase, Nathaniel J. Cosper, David L. Bienvenue, Jacob E. Shokes, Danuta M. Gilner, Takashi Tsukamoto, Robert A. Scott, Richard C. Holz
The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae Is A Dinuclear Metallohydrolase, Nathaniel J. Cosper, David L. Bienvenue, Jacob E. Shokes, Danuta M. Gilner, Takashi Tsukamoto, Robert A. Scott, Richard C. Holz
Richard C. Holz
The Zn K-edge extended X-ray absorption fine structure (EXAFS) spectra, of the dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae have been recorded in the presence of one or two equivalents of Zn(II) (i.e. [Zn_(DapE)] and [ZnZn(DapE)]). The Fourier transforms of the Zn EXAFS are dominated by a peak at ca. 2.0 Å, which can be fit for both [Zn_(DapE)] and [ZnZn(DapE)], assuming ca. 5 (N,O) scatterers at 1.96 and 1.98 Å, respectively. A second-shell feature at ca. 3.34 Å appears in the [ZnZn(DapE)] EXAFS spectrum but is significantly diminished in [Zn_(DapE)]. These data show that DapE contains a dinuclear …
Kinetic And Spectroscopic Analysis Of The Catalytic Role Of H79 In The Methionine Aminopeptidase From Escherichia Coli, Sarah Watterson, Sanghamitra Mitra, Sabina Swierczek, Brian Bennett, Richard Holz
Kinetic And Spectroscopic Analysis Of The Catalytic Role Of H79 In The Methionine Aminopeptidase From Escherichia Coli, Sarah Watterson, Sanghamitra Mitra, Sabina Swierczek, Brian Bennett, Richard Holz
Richard C. Holz
To gain insight into the role of the strictly conserved histidine residue, H79, in the reaction mechanism of the methionyl aminopeptidase from Escherichia coli (EcMetAP-I), the H79A mutated enzyme was prepared. Co(II)-loaded H79A exhibits an overall >7000-fold decrease in specific activity. The almost complete loss of activity is primarily due to a >6000-fold decrease in kcat. Interestingly, the Km value obtained for Co(II)-loaded H79A was approximately half the value observed for wild-type (WT) EcMetAP-I. Consequently, kcat/Km values decreased only 3000-fold. On the other hand, the observed specific activity of Mn(II)-loaded …
Kinetic And Structural Characterization Of Manganese(Ii)-Loaded Methionyl Aminopeptidases, Ventris D'Souza, Sabina Swierczek, Nathaniel Cosper, Lu Meng, Shane Ruebush, Alicja Copik, Robert Scott, Richard Holz
Kinetic And Structural Characterization Of Manganese(Ii)-Loaded Methionyl Aminopeptidases, Ventris D'Souza, Sabina Swierczek, Nathaniel Cosper, Lu Meng, Shane Ruebush, Alicja Copik, Robert Scott, Richard Holz
Richard C. Holz
Manganese(II) activation of the methionyl aminopeptidases from Escherichia coli (EcMetAP-I) and the hyperthermophilic archaeon Pyrococcus furiosus (PfMetAP-II) was investigated. Maximum catalytic activity for both enzymes was obtained with 1 equiv of Mn(II), and the dissociation constants (Kd) for the first metal binding site were found to be 6 ± 0.5 and 1 ± 0.5 μM for EcMetAP-I and PfMetAP-II, respectively. These Kd values were verified by isothermal titration calorimetry (ITC) and found to be 3.0 ± 0.2 and 1.4 ± 0.2 μM for EcMetAP-I and PfMetAP-II, respectively. The hydrolysis of MGMM was measured in triplicate between 25 and 85 °C …
Proton Nmr Spectroscopy As A Probe Of Dinuclear Copper(Ii) Active Sites In Metalloproteins. Characterization Of The Hyperactive Copper(Ii)-Substituted Aminopeptidase From Aeromonas Proteolytica, Richard C. Holz, Brian Bennett, Guanjing Chen, Li-June Ming
Proton Nmr Spectroscopy As A Probe Of Dinuclear Copper(Ii) Active Sites In Metalloproteins. Characterization Of The Hyperactive Copper(Ii)-Substituted Aminopeptidase From Aeromonas Proteolytica, Richard C. Holz, Brian Bennett, Guanjing Chen, Li-June Ming
Richard C. Holz
Proton NMR spectra of the hyperactive Cu(II)-substituted aminopeptidase from Aeromonas proteolytica (AAP) were recorded in both H2O and D2O buffered solution at pH 6.7. Several remarkably sharp, well resolved hyperfine shifted 1H NMR signals were observed in the 70 to −20 ppm chemical shift range. That hyperfine shifted signals were observed is due to spin-coupling of the two Cu(II) ions. Comparison of the spectra recorded in H2O and D2O buffered solutions indicated that the signals at 44.6, 43.3, and 17.7 ppm were solvent exchangeable. The two most strongly downfield shifted signals were assigned to imidazole N−H protons of the two …
Spectroscopic And X-Ray Crystallographic Characterization Of Bestatin Bound To The Aminopeptidase From Aeromonas (Vibrio) Proteolytica, Carin Stamper, David Bienvenue, Brian Bennett, Dagmar Ringe, Gregory Petsko, Richard Holz
Spectroscopic And X-Ray Crystallographic Characterization Of Bestatin Bound To The Aminopeptidase From Aeromonas (Vibrio) Proteolytica, Carin Stamper, David Bienvenue, Brian Bennett, Dagmar Ringe, Gregory Petsko, Richard Holz
Richard C. Holz
Binding of the competitive, slow-binding inhibitor bestatin ([(2S,3R)-3-amino-2-hydroxy-4-phenylbutanoy]-leucine) to the aminopeptidase from Aeromonas proteolytica (AAP) was examined by both spectroscopic and crystallographic methods. Electronic absorption spectra of the catalytically competent [Co_(AAP)], [CoCo(AAP)], and [ZnCo(AAP)] enzymes recorded in the presence of bestatin revealed that both of the divalent metal ions in AAP are involved in binding bestatin. The electron paramagnetic resonance (EPR) spectrum of the [CoCo(AAP)]−bestatin complex exhibited no observable perpendicular- or parallel-mode signal. These data indicate that the two CoII ions in AAP are antiferromagnetically coupled yielding an S = 0 ground state and suggest that a single oxygen atom …
Spectroscopically Distinct Geometrical Isomers In A Single Crystal. Characterization Of The Eight-Coordinate Adducts Of Tris(Dipivaloylmethanato)Lanthanide(Iii) With 2,9-Dimethyl-1,10-Phenanthroline, Richard Holz, Larry Thompson
Spectroscopically Distinct Geometrical Isomers In A Single Crystal. Characterization Of The Eight-Coordinate Adducts Of Tris(Dipivaloylmethanato)Lanthanide(Iii) With 2,9-Dimethyl-1,10-Phenanthroline, Richard Holz, Larry Thompson
Richard C. Holz
Tris(dipivaloylmethanato)(2,9-dimethyl- 1,10-phenanthroline)lanthanide(III), [L n(DPM)3*DMOP], complexes (Ln = La, Eu, Tb, Ho) were synthesized from [Ln(DPM)3] and 2,9-dimethyl- 1,10-phenanthroline in carbon tetrachloride. The Eu3+ and Tb3+ complexes were characterized spectroscopically by infrared and luminescence techniques. The luminescence spectra of the Eu3+ and Tb3+ adducts were recorded at 77 K. For Eu3+, two transitions in the 5Do →FO region are observed which are separated by an unprecedented 35 cm-l. The 5Do →7F1 and 5Do →7F2 spectral regions contain six and eight bands, respectively, indicating two distinct Eu3+ environments. The Eu3+ luminescence spectrum indicates that both Eu3+ environments have local site symmetry C1. …
Epr Studies On The Mono- And Dicobalt(Ii)-Substituted Forms Of The Aminopeptidase From Aeromonas Proteolytica. Insight Into The Catalytic Mechanism Of Dinuclear Hydrolases, Brian Bennett, Richard C. Holz
Epr Studies On The Mono- And Dicobalt(Ii)-Substituted Forms Of The Aminopeptidase From Aeromonas Proteolytica. Insight Into The Catalytic Mechanism Of Dinuclear Hydrolases, Brian Bennett, Richard C. Holz
Richard C. Holz
The structure and function of the prototypical dinuclear hydrolase, namely, the aminopeptidase from Aeromonas proteolytica (AAP), was probed by EPR spectroscopy of the mono- and dicobalt(II)-substituted derivatives. A new systematic protocol for the interpretation of Co(II) EPR spectra is described and the S = 3/2 spin states of the Co(II)-substituted forms of the enzyme have been characterized. This protocol allows the simulation of line shape using theoretically allowed geff values corresponding to an isotropic greal value. In addition, the gross distortion of EPR spectra of high-spin S = 3/2 Co(II) ions has been investigated, and the effects of saturation on …
Synthesis, Molecular Structure And Reactivity Of A New Dicopper(Ii) Benzimidazole Complex With Non-Identical Copper(Ii) Sites, Richard Holz, Feben Gobena
Synthesis, Molecular Structure And Reactivity Of A New Dicopper(Ii) Benzimidazole Complex With Non-Identical Copper(Ii) Sites, Richard Holz, Feben Gobena
Richard C. Holz
The dinucleating ligand ethylene glycol-bis(β-aminoethyl ether) N,N,N′N′-tetrakis[2-(1-ethylbenzimidazoyl)] (EGTB-Et,1) has been synthesized in good yield by the condensation of 1,2-diaminobenzene with ethylene glycol-bis(β-aminoethyl ether) N,N,N′,N′-tetraacetic acid. This ligand was used to synthesize the dinuclear CuII diacetonitrile complex [Cu2(EGTB-Et)(CH3CN)2](ClO4)4·2CH3CN (2). The X-ray structure shows two distinct square pyramidal CuII centres with each CuII ion bound by two benzimidazole nitrogen atoms, one amine nitrogen atom, an ether oxygen atom and one acetinitrile nitrogen atom. The Cu--Cu separation is 5.809A˚. Spectral titration of 2 with sodium acetate or sodium cyanide results in new complexes with terminal acetate or cyano groups. X-band EPR spectra of …
Both Nucleophile And Substrate Bind To The Catalytic Fe(Ii)-Center In The Type-Ii Methionyl Aminopeptidase From Pyrococcus Furiosus, Alicja Copik, Sarah Waterson, Sabina Swierczek, Brian Bennett, Richard Holz
Both Nucleophile And Substrate Bind To The Catalytic Fe(Ii)-Center In The Type-Ii Methionyl Aminopeptidase From Pyrococcus Furiosus, Alicja Copik, Sarah Waterson, Sabina Swierczek, Brian Bennett, Richard Holz
Richard C. Holz
Metalloproteases utilize their active site divalent metal ions to generate a nucleophilic water/hydroxide. For methionine aminopeptidases (MetAPs), the exact location of this nucleophile, as well as of the substrate, with respect to the active site metal ion is unknown. In order to address this issue, we have examined the catalytically competent Fe(II)-loaded form of PfMetAP-II ([Fe(PfMetAP-II)]) in the absence and presence of both nitric oxide (NO) and the substrate-analogue inhibitor butaneboronic acid (BuBA) by kinetic and spectroscopic (EPR, UV−vis) methods. NO binds to [Fe(PfMetAP−II)] with a Kd of 200 μM forming an {FeNO}7 …
Overexpression And Divalent Metal Binding Properties Of The Methionyl Aminopeptidase From Pyrococcus Furiosus, Lu Meng, Shane Ruebush, Ventris D'Souza, Alicja Copik, Susumu Tsunasawa, Richard Holz
Overexpression And Divalent Metal Binding Properties Of The Methionyl Aminopeptidase From Pyrococcus Furiosus, Lu Meng, Shane Ruebush, Ventris D'Souza, Alicja Copik, Susumu Tsunasawa, Richard Holz
Richard C. Holz
The gene encoding for the methionyl aminopeptidase from the hyperthermophilic archaeon Pyrococcus furiosus (PfMetAP-II; EC 3.4.11.18) has been inserted into a pET 27b(+) vector and overexpressed in Escherichia coli. The new expression system resulted in a 5-fold increase in purified enzyme obtained from a 5 L fermentor growth. The as-purified PfMetAP-II enzyme, to which no exogenous metal ions or EDTA was added, was found to have 1.2 equiv of zinc and 0.1 equiv of iron present by ICP-AES analysis. This enzyme had a specific activity of 5 units/mg, a 60-fold decrease from the fully loaded Fe(II) enzymes. When an additional …
The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae Contains Two Active-Site Histidine Residues, Danuta Gillner, David L. Bienvenue, Boguslaw P. Nocek, Andzrej Joachimiak, Vincentos Zachary, Brian Bennett, Richard C. Holz
The Dape-Encoded N-Succinyl-L,L-Diaminopimelic Acid Desuccinylase From Haemophilus Influenzae Contains Two Active-Site Histidine Residues, Danuta Gillner, David L. Bienvenue, Boguslaw P. Nocek, Andzrej Joachimiak, Vincentos Zachary, Brian Bennett, Richard C. Holz
Richard C. Holz
The catalytic and structural properties of the H67A and H349A dapE-encoded N-succinyl-l,l-diaminopimelic acid desuccinylase (DapE) from Haemophilus influenzae were investigated. On the basis of sequence alignment with the carboxypeptidase from Pseudomonas sp. strain RS-16, both H67 and H349 were predicted to be Zn(II) ligands. The H67A DapE enzyme exhibited a decreased catalytic efficiency (180-fold) compared with wild-type (WT) DapE towards N-succinyldiaminopimelic acid. No catalytic activity was observed for H349A under the experimental conditions used. The electronic paramagnetic resonance (EPR) and electronic absorption data indicate that the Co(II) ion bound to H349A-DapE is analogous to that of WT DapE after the …
J-Dependent Curie Or Anti-Curie Behavior Of Proton Nmr Resonances For Antiferromagnetically Coupled Dinuclear Copper(Ii) Complexes, Richard Holz, Julie Brink, Rachel Rose
J-Dependent Curie Or Anti-Curie Behavior Of Proton Nmr Resonances For Antiferromagnetically Coupled Dinuclear Copper(Ii) Complexes, Richard Holz, Julie Brink, Rachel Rose
Richard C. Holz
No abstract provided.
Inhibition Of The Aminopeptidase From Aeromonas Proteolytica By L-Leucinephosphonic Acid. Spectroscopic And Crystallographic Characterization Of The Transition State Of Peptide Hydrolysis, Carin Stamper, Brian Bennett, Tanya Edwards, Richard Holz, Dagmar Ringe, Gregory Petsko
Inhibition Of The Aminopeptidase From Aeromonas Proteolytica By L-Leucinephosphonic Acid. Spectroscopic And Crystallographic Characterization Of The Transition State Of Peptide Hydrolysis, Carin Stamper, Brian Bennett, Tanya Edwards, Richard Holz, Dagmar Ringe, Gregory Petsko
Richard C. Holz
The nature of the interaction of the transition-state analogue inhibitor l-leucinephosphonic acid (LPA) with the leucine aminopeptidase from Aeromonas proteolytica (AAP) was investigated. LPA was shown to be a competitive inhibitor at pH 8.0 with a Ki of 6.6 μM. Electronic absorption spectra, recorded at pH 7.5 of [CoCo(AAP)], [CoZn(AAP)], and [ZnCo(AAP)] upon addition of LPA suggest that LPA interacts with both metal ions in the dinuclear active site. EPR studies on the Co(II)-substituted forms of AAP revealed that the environments of the Co(II) ions in both [CoZn(AAP)] and [ZnCo(AAP)] become highly asymmetric and constrained upon the addition of …