Structural Studies of Herpesvirus Proteases Pp.333-342
Renu
Batra, Reza Khayat and Liang Tong
Atomic Force Microscopy Investigation of Ribonuclease A Pp.343-347
Luciano
Paulino da Silva
Nucleolin
Binds to a cis-Active Region in the Human Desmocollin 2 Promoter Pp.349-356
Paresh
L. Gadhavi, Steven Howell, Anthony I. Magee and Roger S. Buxton
Hydroxyapatite Elution Behavior of Human Nucleotide Excision Repair Protein XPA and Fragments of XPA Pp.357-365
Garry
W. Buchko and Michael A. Kennedy
Isolation and Identification of Two Membrane Proteins Binding to Transfer RNA Pp.367-374
Quan-Sheng
Liu, Jian-Hua Liu, De-Sheng Jiang, You-Xin Jin and De-Bao Wang
Detection and Purification of Insecticidal Peptides from Scorpions and Spiders: A Rapid Method for Their Isolation from Their Crude Venoms Using MALDI-TOF-MS Analysis Pp.375-383
Gerardo
Corzo, Elba Villegas, Hon-Seok Lee and Terumi Nakajima
Isolation and Structural Characterization of a Peptide from the Venom of Scorpion with Toxicity Towards Invertebrates and Vertebrates Pp.385-393
Gerardo Corzo, Elba Villegas and Terumi Nakajima
Effect of
Phospholipid Membrane on Conformation of Retro-Sequence Amyloid- b-Peptide 35-25 and Normal Amyloid-b-Peptide 25-35 Pp.395-398
Masato Kodaka
Macromolecular
Properties of Diocleinae Lectins Pp.399-406
Márcio
V. Ramos, Leila M. Beltramini and Renato A. Moreira
Inhibition of Fungal Aldose Reductase Pp.407-412
Ya-Jun
Zheng, Yong Tao, Wei Zhang and Douglas B. Jordan
Cross-Neutralizing Human Monoclonal Anti-HIV-1 Antibody 2F5: Preparation and Crystallographic Analysis of the Free and Epitope-Complexed Forms of its Fab' Fragment Pp.413-418
Steve
Bryson, Annie Cunningham, Jason Ho, Rosemary C. Hynes, David E. Isenman, Brian
H. Barber, Renate Kunert, Hermann Katinger, Michel Klein and Emil F. Pai
Crystallization and Preliminary X-Ray Crystallographic Analysis of bHLH Domain of Aryl Hydrocarbon Receptor Nucleus Translocator Pp.419-422
Ji-Joon Song, Mun-Kyoung Kim, Eunseon Hur, Hyunsung Park and Soo Hyun Eom
Abstracts
[Back to top] Structural Studies
of Herpesvirus Proteases
Renu Batra, Reza Khayat and Liang Tong
Structural studies of
herpesvirus proteases establish that they belong to a new class of serine
proteases and contain a novel Ser-His-His catalytic triad. Peptidomimetic
inhibitors bind to the protease by forming an anti-parallel b-sheet with the
enzyme. There are large conformational changes in the protease upon inhibitor
binding, indicating that the protease is an induced-fit enzyme. Further studies
are needed to understand the molecular basis for the dimerization requirement
of the protease.
[Back to top] Atomic Force Microscopy Investigation of
Ribonuclease A
Luciano Paulino da Silva
Ribonuclease
A (RNase A) molecules have been adsorbed onto mica surfaces from aqueous
solution at pH 7.4. Atomic force microscopy (AFM) images of native RNase A show
aggregates of monomers each having a diameter of 9 nm. This is consistent with
a globular unit the size of which would be substantially larger than that
expected for a 13-kDa protein. These experiment suggest that the RNase A
oligomer subunits exist as a multimeric complex with the 13-kDa protein.
[Back to top]
Nucleolin Binds to a cis-Active
Region in the Human Desmocollin 2 Promoter
Paresh L. Gadhavi, Steven Howell, Anthony I. Magee and Roger S. Buxton
Affinity chromatography using oligo DNA from a cis-active region (-453/-389) of the human DSC2 promoter was used to isolate a protein from human CaCo-2 epithelial cells expressing DSC2. The protein was SDS-PAGE purified, trypsinized and analyzed by MALDI-TOF mass spectrometry. Peptide mass fingerprinting identified the protein as nucleolin, a ubiquitously expressed protein involved in gene regulation
[Back to top] Hydroxyapatite Elution Behavior of Human
Nucleotide Excision Repair Protein XPA and Fragments of XPA
Garry W. Buchko and Michael A. Kennedy
The hydroxyapatite elution behavior of 3 fragments of human XPA (M98-F219 = XPA-MBD; M59-F219 = XPA-EM; M59-M273 = XPA-DN58) and full-length human XPA containing a 6-residue, N-terminal histidine tag (h6-XPA) are described. With increasing concentration of potassium phosphate the proteins elute in the order: XPA-EM, XPA-MBD, XPA-DN58, and h6-XPA. If hydroxyapatite affinity is related to DNA-binding affinity, then h6-XPA and XPA-DN58 may bind DNA more tightly than XPA-MBD and XPA-EM.
[Back to top] Isolation and Identification of Two Membrane
Proteins Binding to Transfer RNA
Quan-Sheng
Liu, Jian-Hua Liu, De-Sheng Jiang, You-Xin Jin and De-Bao Wang
Two major tRNA binding proteins were obtained by biochemical method from yeast crude extracts of total membrane proteins. They were further confirmed to be tRNA specific-binding proteins by gel mobility shift assay. N-terminal sequence assays showed that they were glyceraldehyde-3-phosphate dehydranase (GAPDH) and 3-phosphate glycerate kinase (PGK1), respectively.
[Back to top] Detection and Purification of Insecticidal Peptides from Scorpions and Spiders A Rapid Method for Their Isolation from Their Crude Venoms Using MALDI-TOF-MS Analysis
Gerardo
Corzo, Elba Villegas, Hon-Seok Lee and Terumi Nakajima
The insecticidal neurotoxins AaIT, LqhIT2, and m-agaIV from venom of Arachnida were rapidly re-purified from their respective crude venoms using mass spectrometry analyses and two high pressure chromatographic steps. The purity of each toxin was confirmed by capillary zone electrophoresis. Each toxin was identified by mass spectrometry and their biological activity confirmed by bioassays against tobacco cutworms and mice. In addition, AaIT and LqhIT2 were partially sequenced using post-source decay mass spectrometry analysis. The spider neurotoxin m-agaIV was identified from other isomassic peptide by fingerprint mass analysis of its tryptic fragments.
[Back to top] Isolation and Structural Characterization
of a Peptide from the Venom of Scorpion with Toxicity Towards Invertebrates and
Vertebrates
Gerardo Corzo, Elba Villegas and Terumi Nakajima
Using a methodology based on HPLC fractionation, mass
spectrometry analysis, and microinjection guide-bioassays, a peptide with
toxicity to both insects and mammals was isolated from the venom of the
scorpion Leiurus quinquestriatus hebraeus, and named LqhIV (after LqhII and
LqhIII[1]). The primary sequence, circular dichroism spectrum
and three-dimensional structure simulation of LqhIV indicate that it contains
similar structure to that of the group of alpha scorpion toxins.
[Back to top] Effect of Phospholipid Membrane on
Conformation of Retro-Sequence Amyloid b-Peptide
35-25 and Normal Amyloid-b-Peptide
25-35
Masato
Kodaka
Negatively-charged phospholipid membrane does not affect the random coil conformation of a non-neurotoxic amyloid-b-peptide having a retro-sequence 35-25 (Ab 35-25), whereas it changes the conformation of a neurotoxic normal-sequence peptide Ab 25-35 from a random coil to a b-sheet-like structure under the same condition. The formation of the ordered structure seems to be closely related to the expression of neurotoxicity.
[Back to top] Macromolecular Properties of Diocleinae
Lectins
Márcio V. Ramos, Leila M. Beltramini and Renato A. Moreira
Structural properties of six
closely related lectins isolated from plants belonging from the legume group
Diocleinae were studied in solution. Affinity and gel filtration chromatography
of the lectins at different pH conditions demonstrated that the lectins diverge
in affinity for a specific matrix mainly at the pH range 4.0 up to 6.0 and that
at neutral pH they tend to form dimers. Analysis of circular dichroism and
fluorescence emission spectra stressed the differences of the lectin from
Cratylia floribunda seeds from that of Canavalia brasiliensis, C. ensisformis,
C. maritima, Diolcea grandiflora and D.virgata. According to the results, these
proteins constitute a excellent model to study fine protein structure/function
relationships.
[Back to top] Inhibition of Fungal Aldose Reductase
Ya-Jun
Zheng, Yong Tao, Wei Zhang and Douglas B. Jordan
Aldose reductase (AR) was
cloned from the fungal pathogen, Magnaporthe grisea, expressed in Escherichia
coli and purified to homogeneity. An experimental fungicide inhibits the M.
grisea AR uncompetitively with respect to its aldehyde substrate with a Ki
of 130 nM, the potency suggesting that AR is a biochemical target of the
fungicide. The M. grisea AR has a high kcat and large differences in
substrate specificities.
[Back to top] Cross-Neutralizing Human Monoclonal Anti-HIV-1 Antibody 2F5: Preparation and Crystallographic Analysis of the Free and Epitope-Complexed Forms of its Fab' Fragment
Steve
Bryson, Annie Cunningham, Jason Ho, Rosemary C. Hynes, David E. Isenman, Brian
H. Barber, Renate Kunert, Hermann Katinger, Michel Klein and Emil F. Pai
The human monoclonal antibody
2F5 is a potent neutralizer of most clades of HIV-1 and possesses protective
effects against viral transmission. It recognizes the linear epitope ELDKWAS of
the viral envelope protein gp41. As structural information about epitope
recognition may help to develop an HIV-1 vaccine we initiated crystallographic
analyses of mAb 2F5 and its epitope complex. We now report the preparation of
the corresponding Fab’ fragments, complexation with the epitope peptide, and
crystallization of free mAb 2F5 Fab’ as well as the peptide complex. Both
crystal forms are well suited for high-resolution structural analysis.
[Back to top] Crystallization and Preliminary X-Ray
Crystallographic Analysis of bHLH Domain of Aryl Hydrocarbon Receptor Nucleus
Translocator
Ji-Joon
Song, Mun-Kyoung Kim, Eunseon Hur, Hyunsung Park and Soo Hyun Eom
The basic-helix-loop-helix
(bHLH) DNA binding domain of Aryl hydrocarbon receptor nucleus translocator
(Arnt) was purified and crystallized in glutathione S transferase (GST) fusion
form. The tetragonal crystal was grown in ammonium sulfate condition and
diffracted up to 2.2 Ĺ. The space group was determined as P43212 with a = b =
92.2, c = 57.6 Ĺ unit cell parameters.