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Protein & Peptide Letters
ISSN: 0929-8665
Protein & Peptide Letters
Volume 14, Number 1, 2007
Contents
Regular Papers
Neurotoxic Prion Protein (PrP) Fragment 106-126 Requires the
N-Terminal Half of the Hydrophobic Region of PrP in the PrP-Deficient
Neuronal Cell Line Pp. 1-6
A. Sakudo, I. Nakamura, D.-C. Lee, K. Saeki, K. Ikuta
and T. Onodera
[Abstract] [Full
Text Article]
Expression and Antibody Preparation of POU Transcription
Factor qBrn-1 Pp. 7-11
L. Lan, M. Liu, Y. Liu and R. He
[Abstract] [Full
Text Article]
Spectroscopic Studies of the Effects of Glycation
of Human Serum Albumin on L-Trp Binding Pp. 13-18
A. Barzegar, A.A. Moosavi-Movahedi, N. Sattarahmady, M.A.
Hosseinpour-Faizi, M. Aminbakhsh, F. Ahmad, A.A. Saboury,
M.R. Ganjali and P. Norouzi
[Abstract] [Full
Text Article]
Stability of Synthetic Exendin-4 in Human Plasma In
Vitro Pp. 19-25
J. Chen, L. Yu, L. Wang, X. Fang, L. Li and W. Li
[Abstract] [Full
Text Article]
Residues Around the Catalytic Pocket of DdsA Are Important
for Isoprenoids Chain Elongation Pp. 27-32
X. Liu, Q. Yuan and H. Zhang
[Abstract] [Full
Text Article]
Binding Sites on Laminin Receptors as Components for
Antibiotics Pp. 33-36
N. Kobayashi and T. Yoshida
[Abstract] [Full
Text Article]
Discrimination of Outer Membrane Proteins by a New
Measure of Information Discrepancy Pp. 37-44
Z. Wu, E. Feng, Y. Wang and L. Chen
[Abstract] [Full
Text Article]
Preliminary Estimation of Rotary Torque Produced by
Proton-Motive Force in Fully Functional F0F1-ATPase
Pp. 45-50
X. Liu, Y. Cui, C. Chen, B. Lai, J. Yue and Z. Zhang
[Abstract] [Full
Text Article]
Topological Exploration of Cyclic Endomorphin-1 Analogues,
Structurally Defined Models for Investigating the Bioactive
Conformation of MOR Agonists Pp. 51-56
L. Gentilucci, A. Tolomelli and F. Squassabia
[Abstract] [Full
Text Article]
Tool Developments for Structure-Function Studies of
Host Defense Peptides Pp. 57-69
G. Wang
[Abstract] [Full
Text Article]
Characterization of a Lectin from Gonatanthus
pumilus D. Don Having Anti-Proliferative Effect Against
Human Cancer Cell Lines Pp. 71-78
V. Dhuna, S.S. Kamboj, A. Kaur, A.K. Saxena, S.V. Bhide,
Shanmugavel and J. Singh
[Abstract] [Full
Text Article]
Unexpected Altered Specificity Is Responsible for
St. Louis Encephalitis Virus Recombinant Protease
Autoproteolysis Pp. 79-82
B.A.M. Pastorino, C.N. Peyrefitte, M. Grandadam, R. Lebrun,
D. Moinier, D. Rolland, H.J. Tolou and M. Bessaud
[Abstract] [Full
Text Article]
Aβ
Produced as a Fusion to Maltose Binding Protein Can Be Readily
Purified and Stably Associates with Copper and Zinc
Pp. 83-86
J. Caine, I. Volitakis, R. Cherny, J. Varghese and I.
Macreadie
[Abstract] [Full
Text Article]
Large Scale Preparation of the Mammalian High Mobility
Group Protein A2 for Biophysical Studies Pp. 87-91
T. Cui, S. Joynt, V. Morillo, M. Baez, Z. Hua, X. Wang
and F. Leng
[Abstract] [Full
Text Article]
Crystallization Reports
Crystallization and Preliminary Diffraction Studies
of Porcine Pancreatic Elastase in Complex with a Novel Inhibitor
Pp. 93-95
T.F. Oliveira, J. Mulchande, R. Moreira, J. Iley and M.
Archer
[Abstract] [Full
Text Article]
Crystallization and Preliminary X-Ray Diffraction
Analysis of PD-L4, a Ribosome Inactivating Protein from Phytolacca
dioica L. leaves Pp. 97-100
A. Ruggiero, A. Chambery, A. Di Maro, M. Pisante, A. Parente
and R. Berisio
[Abstract] [Full
Text Article]
Abstracts
[Back to top]
Neurotoxic Prion Protein (PrP) Fragment 106-126 Requires
the N-Terminal Half of the Hydrophobic Region of PrP in the
PrP-Deficient Neuronal Cell Line
A. Sakudo, I. Nakamura, D.-C. Lee, K. Saeki, K. Ikuta
and T. Onodera
[Full
Text Article]
The cytotoxicity of aged PrP(106-126) was examined using
an immortalized prion protein (PrP) gene-defcient neuronal
cell line. The N-terminal half of the hydrophobic region (HR)
but not the octapeptide repeat (OR) of PrP was required for
aged PrP(106-126) neurotoxicity, suggesting that neurotoxic
signals of aged PrP(106-126) are mediated by this region.
[Back to top]
Expression and Antibody Preparation of POU Transcription
Factor qBrn-1
L. Lan, M. Liu, Y. Liu and R. He
[Full
Text Article]
The transcription factor Brn-1, which belongs to POU-domain
family, has been shown to play critical roles in the development
of the nervous system. A cDNA clone coding for a quail Brn-1
homologue, qBrn-1, was isolated. To investigate whether this
gene plays a role in the development of the quail nervous
system, an anti-N-terminal peptide anti-body was prepared.
The coding region for amino acids 1-79 (the N-terminal domain
of qBrn-1) was subcloned into Trx fusion expression vector
pET32c and introduced into the Escherichia coli Origami(DE3)
cells for efficient expression. After purification, Trx-fused
polypeptides, called Trx-qBrn-1N, were used to immunize the
rabbits to prepare polyclonal antibodies against qBrn-1. The
produced and purified antiserum showed specificity not only
to the in vitro expressed qBrn-1, but also to the
natural qBrn-1 in tissues. Immunolabeling on sections by the
anti-qBrn-1 serum showed that qBrn-1 was specifically expressed
in the developing spinal cord and kidney. This suggests that
qBrn-1 may play some roles in the development of avian nervous
system and kidney, and the preparation of anti-qBrn-1 polyclonal
antibody will facilitate further detection of, and functional
study on, qBrn-1 both in vivo and in vitro.
[Back to top]
Spectroscopic Studies of the Effects of Glycation
of Human Serum Albumin on L-Trp Binding
A. Barzegar, A.A. Moosavi-Movahedi, N. Sattarahmady, M.A.
Hosseinpour-Faizi, M. Aminbakhsh, F. Ahmad, A.A. Saboury,
M.R. Ganjali and P. Norouzi
[Full
Text Article]
Modification of proteins by nonenzymatic glycation is one
of the underlying factors that contribute to the development
of the complications of diabetes. Human serum albumin (HSA)
is one of the major targets of interaction with glucose through
the Maillard reaction. The effects of 1 and 5 mg/ml glucose
concentrations, which are consistent with blood glucose levels
found in diabetic patients, on human serum albumin were studied
by circular dichroism and fluorescence spectroscopy in sodium
phosphate buffer, pH 7.4. Partial denaturation and changes
in the structural integrity of HSA are caused by glycation
at lower (1 mg/ml) and higher (5 mg/ml) concentrations of
glucose. To study the relationship between structure and function,
we investigated the interaction of L-tryptophan (L-Trp) with
glycated and non-glycated HSA. The results showed that L-Trp,
as the only free amino acid that substantially binds to HSA,
has a lower affinity for the glycated form (especially at
low concentrations of glucose) than for non-glycated HSA.
[Back to top]
Stability of Synthetic Exendin-4 in Human Plasma In
Vitro
J. Chen, L. Yu, L. Wang, X. Fang, L. Li and W. Li
[Full
Text Article]
Sites of exendin-4 that that are relatively susceptible to
degradation in plasma were identified with the aim of providing
information for designing new exendin-4 analogues. The stability
of exendin-4 in human plasma was evaluated in vitro.
The results showed that the peptide was slowly degraded with
a half-life of 9.57 h and the principal cleavage sites are
between Thr5 and Phe6, Phe6 and Thr7, and Thr7 and Ser8 of
the N-terminus region of exendin-4.
[Back to top]
Residues Around the Catalytic Pocket of DdsA Are Important
for Isoprenoids Chain Elongation
X. Liu, Q. Yuan and H. Zhang
[Full
Text Article]
Nineteen mutants of decaprenyl diphosphate synthase from Gluconobacter
suboxydans were generated and their production of ubiquinones
(UQs) was compared to that of the wild type protein. The accessible
surface area and the polarity of amino acid around the catalytic
pocket wield remarkable influences on the isoprenoid chain
elongation of UQs.
[Back to top]
Binding Sites on Laminin Receptors as Components for
Antibiotics
N. Kobayashi and T. Yoshida
[Full
Text Article]
Bacteria use the receptor-adhesion-like interaction between
laminin and the laminin receptor in the process of infection.
We determined that bacteria do not interact with the receptor-binding
site on laminin which could be expected for the bacterial
laminin receptor. Rather, binding occurs via the laminin-binding
site on the 67-kDa laminin receptor, which has a function
similar to the one the bacterial laminin receptor possesses.
This finding has implications for the effective use of antimicrobial
peptides.
[Back to top]
Discrimination of Outer Membrane Proteins by a New
Measure of Information Discrepancy
Z. Wu, E. Feng, Y. Wang and L. Chen
[Full
Text Article]
Discriminating outer membrane proteins for globular proteins
(GPs) and other types of membrane proteins from genomic sequences
is an important and hot topic. In this paper, a measure based
on information discrepancy is proposed and applied to the
discrimination of outer membrane proteins. It differs from
previous methods which are based on amino acid composition.
Our approach focuses on the comparison of subsequence distributions
and takes into account the effect of residue order in protein
primary structures. As a result, the new approach outperforms
all previous methods on the same benchmark datasets. In particular,
we show that the proposed approach has correctly identified
the outer membrane proteins at an accuracy of 99% for the
training set of 337 proteins and has correctly excluded the
GPs at an accuracy of 86% in a non-redundant dataset of 668
proteins. Furthermore, this method is able to correctly exclude
α-helical
membrane proteins at an accuracy of 100%.
[Back to top]
Preliminary Estimation of Rotary Torque Produced by
Proton-Motive Force in Fully Functional F0F1-ATPase
X. Liu, Y. Cui, C. Chen, B. Lai, J. Yue and Z. Zhang
[Full
Text Article]
F0F1-ATPase is a rotary molecular motor.
It is well known that the rotary torque is generated by ATP
hydrolysis in F1 but little is known about how
it produces the proton-motive force (PMF) in F0.
Here a cross-linking approach was used to estimate the rotary
torque produced by PMF. Three mutant E. coli strains
were used in this study: SWM92 (δW28L
F0F1, as control), MM10 (αP280CγA285C
F0F1) and PP2 (αA334C/γL262C
F0F1). The oxidized inner membranes
from mutant MM10 having a disulfide bridge in the top of γ
subunit exhibited good ATP synthesis activity, while the oxidized
PP2 inner membranes having a disulfide bridge in the middle
of γ
subunit synthesized ATP very poorly. We conclude that the
rotary torque generated by PMF is sufficient to uncoil the
α-helix
in the top of γ
subunit (MM10) and to overcome the Ramachandran activation
barriers (25-30kJ/mol, i.e. about 40-50pN•nm), but cannot
cleave the disulfide bond in the middle of the γ
subunit (200 kJ/mol, i.e. 330pN•nm) (PP2). Consequently
a preliminary estimation is that the rotary torque generated
by PMF in the fully functional F0F1
motor is greater than 40-50pN•nm but less than 330pN•nm.
[Back to top]
Topological Exploration of Cyclic Endomorphin-1 Analogues,
Structurally Defined Models for Investigating the Bioactive
Conformation of MOR Agonists
L. Gentilucci, A. Tolomelli and F. Squassabia
[Full
Text Article]
Although there have been several reports on the conformational
analysis of endomorphin-1 (YPWF-NH2) and related
MOR (μ-opioid
receptor) agonists, a definitive, convincing model of the
biologically active structure is not yet available. We recently
reported the synthesis and pharmacological characterization
of the atypical endomorphin-analogue agonist c[YpwFG].
In this paper we discuss the conformational analysis of c[YpwFG]
in comparison to its epimers, for investigating the topological
features responsible for ligand recognition and receptor activation,
and the role of the different pharmacophores.
[Back to top]
Tool Developments for Structure-Function Studies of
Host Defense Peptides
G. Wang
[Full
Text Article]
Antimicrobial peptides, or host defense peptides, are universal
signaling and effector molecules in host defense and innate
immunity. This article highlights various tools developed
for cathelicidins and defensins, ranging from peptide identification,
production, and structural biology, including the eight databases
for antimicrobial peptides. Novel peptides can be identified
from natural sources at both gene and protein levels. Solid-phase
synthesis and bacterial expression are the two important methods
for peptide production. Three-dimensional structures of antimicrobial
peptides, primarily determined by solution NMR techniques,
are essential for an in-depth understanding of the mode of
action. The introduction of octanoyl phosphatidylglycerol
as a bacterial membrane-mimetic model provides new insights
into peptide-lipid interactions. The incorporation of structure
and activity data into the antimicrobial peptide database
(http://aps.unmc.edu/AP/main.html) will lead to an integrated
understanding of these peptides via structural bioinformatics.
[Back to top]
Characterization of a Lectin from Gonatanthus
pumilus D. Don Having Anti-Proliferative Effect Against
Human Cancer Cell Lines
V. Dhuna, S.S. Kamboj, A. Kaur, A.K. Saxena, S.V. Bhide,
Shanmugavel and J. Singh
[Full
Text Article]
A monocot araceous lectin from tubers of Gonatanthus pumilus
(GPL) was earlier purified in our laboratory and reported
as T-cell mitogen having homotetrameric structure with subunit
molecular mass of 13 kDa. Besides asialofetuin as reported
earlier, in the present study it was also inhibited by N-acetyl-D-lactosamine
but was non-reactive towards mannose or its derivatives. The
lectin is rich in acidic amino acids and cysteine is completely
absent. Chemical modification of GPL revealed requirement
of tryptophan and tyrosine for lectin sugar interaction. The
secondary structure content of GPL, as estimated with CD spectrum
in K2D programme, has 37% α-helix,
26% β-sheet
and 38% random contributions. Fluorescence spectrum of the
lectin solution at 280 nm was typical for tryptophan residues
buried inside the protein. Lectin activity decreased when
treated with denaturants like guanidine-HCl, urea and thiourea.
GPL inhibited the growth of three plant pathogenic fungi namely
Colletotrichum lindemuthianum, Fusarium oxysporum
and Botrytis cinerea. Out of 11 human cancer cell
lines tested, GPL significantly inhibited proliferation of
five lines viz. Colo-205, IMR-32, HCT-15, SK-N-SH and HT-29.
[Back to top]
Unexpected Altered Specificity Is Responsible for
St. Louis Encephalitis Virus Recombinant Protease
Autoproteolysis
B.A.M. Pastorino, C.N. Peyrefitte, M. Grandadam, R. Lebrun,
D. Moinier, D. Rolland, H.J. Tolou and M. Bessaud
[Full
Text Article]
We report herein the study of the cleavage fragments generated
by autoproteolysis of the St. Louis encephalitis virus
recombinant protease. The cleavage sites leading to truncated
forms were identified by microsequencing, which revealed an
unexpected altered specificity of the recombinant proteinase
towards unusual sequences.
[Back to top]
Aβ
Produced as a Fusion to Maltose Binding Protein Can Be Readily
Purified and Stably Associates with Copper and Zinc
J. Caine, I. Volitakis, R. Cherny, J. Varghese and I.
Macreadie
[Full
Text Article]
The 42 amino acid Alzheimer’s Aβ
peptide has been produced in E. coli as a soluble
fusion to maltose binding protein (MBP). Affinity purification
on amylose columns of MBP-Aβ
and MBP led to the recovery of proteins at purities that were
suited for physicochemical analyses. MBP-Aβ
was able to bind approximately 2 mole equivalents of copper
or 4 mole equivalents of zinc, while MBP alone bound negligible
amounts of zinc or copper. We conclude that Aβ
can bind 2 copper or 4 zinc ions in its fusion format. Because
MBP-Aβ
is a convenient protein to work with, this system is well
suited for further studies on the structure of Aβ
and its interactions with metals.
[Back to top]
Large Scale Preparation of the Mammalian High Mobility
Group Protein A2 for Biophysical Studies
T. Cui, S. Joynt, V. Morillo, M. Baez, Z. Hua, X. Wang
and F. Leng
[Full
Text Article]
Due to asymmetrical charge distribution of the mammalian high
mobility group protein A2 (HMGA2), which makes HMGA2 bind
to both cation- and anion-exchange columns, we developed a
rapid procedure for purifying HMGA2 in the milligram range.
This purification procedure greatly facilitated biophysical
studies, which require large amounts of the protein.
[Back to top]
Crystallization and Preliminary Diffraction Studies
of Porcine Pancreatic Elastase in Complex with a Novel Inhibitor
T.F. Oliveira, J. Mulchande, R. Moreira, J. Iley and M.
Archer
[Full
Text Article]
Porcine pancreatic elastase (PPE) was crystallized in complex
with a novel inhibitor at pH 5 and X-ray diffraction data
were collected at a synchrotron source to 1.66 Å. Crystals
belong to the orthorhombic space group P212121,
with unit cell parameters a = 50.25 Å, b = 57.94 Å
and c = 74.69 Å. PPE is often used as model for drug
target, due to its structural homology with the important
therapeutic target human leukocyte elastase (HLE). Elastase
is a serine protease that belongs to the chymotrypsin family,
which has the ability to degrade elastin, an important component
in connective tissues. Excessive elastin proteolysis leads
to a number of pathological diseases.
[Back to top]
Crystallization and Preliminary X-Ray Diffraction
Analysis of PD-L4, a Ribosome Inactivating Protein from Phytolacca
dioica L. leaves
A. Ruggiero, A. Chambery, A. Di Maro, M. Pisante, A. Parente
and R. Berisio
[Full
Text Article]
PD-L4, a type 1 ribosome inactivating protein from Phytolacca
dioica leaves, has been successfully crystallized using
vapour diffusion methods and PEG 4000 as a precipitant agent.
In addition, crystals of a PD-L4 mutant, which has been recently
observed to have a lower polynucleotide-adenosine glycosidase
activity on DNA, rRNA and poly (A) substrates, have been obtained.
To gather information on PD-L4 reaction mechanism both forms
have been co-crystallized with adenine, the major product
of their catalytic reaction. Diffraction patterns extend to
atomic resolution and crystals belong to the orthorhombic
P212121 space group, with
one molecule in the asymmetric unit. Structure determination
has been achieved using molecular replacement; preliminary
electron density maps have clearly given evidence of adenine
binding.
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