| Protein
& Peptide Letters
ISSN: 0929-8665
Protein & Peptide Letters
Volume 15, Number 7, 2008
Contents
Regular Papers
Oligomerization and Aggregation of Bovine Pancreatic
Ribonuclease A: Backbone Hydration Probed by Infrared Band
Shift Pp. 650-657
J. Zhang and Y.-B. Yan
[Abstract]
Effects of Entomopathogenic Bacterium Photorhabdus
temperata Infection on the Digestive Enzymes of Diatraea
saccharalis (Lepidoptera: Crambidae) Larvae Pp.
658-662
C.N.B. Carneiro, R.A. DaMatta, R.I. Samuels
and C.P. Silva
[Abstract]
Albumin Competitively Inhibits Glycation
of Less Abundant Proteins Pp. 663-667
H.S. Bhonsle, S.K. Singh, G. Srivastava, R. Boppana
and M.J. Kulkarni
[Abstract]
Multiple Ligands in Opioid Research
Pp. 668-682
S. Ballet, M. Pietsch and A.D.
Abell
[Abstract]
Opioid Peptides and Innate Immune Response
in Mollusc Pp. 683-686
D.-W. Liu
[Abstract]
Efficient Expression of Membrane-Bound
Water Channel Protein (Aquaporin Z) in Escherichia coli
Pp. 687-691
J. Lian, X. Fang, J. Cai, Q. Chen, Q. Zheng,
L. Kai and Z. Xu
[Abstract]
A Protein Interaction Network Analysis
for Yeast Integral Membrane Protein Pp.
692-699
M.-G. Shi, D.-S. Huang and X.-L. Li
[Abstract]
A New Family of Small (4kDa) Neurotoxins
from the Venoms of Spiders of the Genus Phoneutria Pp.700-708
A.D. Lúcio, F.V. Campos, M. Richardson,
M.N. Cordeiro, M.S.C. Mazzoni, M.E. de Lima, A.M.C. Pimenta,
M.P. Bemquerer, S.G. Figueiredo, P.C. Gomes and P.S.L.
Beirão
[Abstract]
Crystal Structure of SCO6571 from Streptomyces
coelicolor A3(2) Pp. 709-712
P. Begum, N. Sakai, T. Hayashi, Y.-G. Gao,
T. Tamura, N. Watanabe, M. Yao and I. Tanaka
[Abstract]
Structural and Spectroscopic Elucidation
of Tetrapetide Glycyl-L-Prolyl-Glycyl-Glycine
and Its Hydrogensquarate Pp. 713-718
T.M. Kolev
[Abstract]
Yellow Lupine Cyclophilin Interacts with
Nucleic Acids Pp. 719-723
K. Nuc, K. Lesniewicz, P. Nuc and
R. Slomski
[Abstract]
Purification, Biochemical and Functional
Characterization of Miliin, a New Thiol-Dependent Serine Protease
Isolated from the Latex of Euphorbia milii Pp.
724-730
L.P. Moro, M.T. Murakami, H. Cabral, A.
Vidotto, E.H. Tajara, R.K. Arni, L. Juliano and G.O.
Bonilla-Rodriguez
[Abstract]
Three Sampling Strategies to Predict
Mutations in H5N1 Hemagglutitins from Influenza A Virus
Pp. 731-738
G. Wu and S. Yan
[Abstract]
Predicting Subcellular Localization of
Mycobacterial Proteins by Using Chou’s Pseudo Amino
Acid Composition Pp. 739-744
H. Lin, H. Ding, F.-B. Guo, A.-Y. Zhang
and J. Huang
[Abstract]
Construction, Expression, Purification
and Immunology Effect of an Anti-Atherosclerosis Chimeric
Enzyme Vaccine in Escherichia coli Pp. 745-752
S. Yunxiao, L. Zhufang, Y. Xin, L. Jun,
X. Qiyan, Q. Gaofu, C. Rongyue, W. Jie, L. Taiming, F. Hao
and L. Jingjing
[Abstract]
Crystallization Report
Preliminary Structural Studies on MPN423 Expressed from an
Orthologous ORFan of Mycoplasma pneumoniae Pp.
753-755
D.H. Shin
[Abstract]
Abstracts
[Back to top]
Oligomerization and Aggregation of Bovine Pancreatic
Ribonuclease A: Backbone Hydration Probed by Infrared Band
Shift
J. Zhang and Y.-B. Yan
Protein hydration plays a crucial role in almost all aspects
of biomolecular processes. In this research, we studied the
hydration/dehydration-induced infrared amide I band-shift
by using poly-L-lysine and bovine pancreas ribonuclease A
as model polypeptides. It was found that a 1-4 cm-1
shift could be clearly distinguished for all regular secondary
structures during protein thermal unfolding. This shift was
proven to be due to backbone hydration but not from experimental
error, temperature effect or possible incomplete hydrogen/deuterium
exchange of the samples. Moreover, we also found that protein
aggregation was closely associated with the backbone hydration/dehydration
status of proteins. In conditions favoring aggregation, a
significant shift to a higher wavenumber of the band from
the intermolecular β-sheet
structures in aggregates was observed. The present study suggested
that the changes of the amounts of regular secondary structures
could be monitored by the intensity changes, while the changes
of the hydration status could be monitored by the shift of
the infrared bands.
[Back to top]
Effects of Entomopathogenic Bacterium Photorhabdus
temperata Infection on the Digestive Enzymes of Diatraea
saccharalis (Lepidoptera: Crambidae) Larvae
C.N.B. Carneiro, R.A. DaMatta, R.I. Samuels
and C.P. Silva
This study investigated the effects of Photorhabdus temperata
infection on the activities of digestive enzymes of the sugarcane
stalk borer Diatraea saccharalis. Non-infected D.
saccharalis larvae present a major α-amylase,
several proteinases, three sucrose hydrolases and two α-glucosidases
in their midgut. Analysis of these hydrolases by electrophoresis
and “in gel” assays showed that the activities
of all enzymes decreased following infection, with an initial
decline observed 12 h after infection. The activities of α-glucosidases
decreased by 50% twelve hours after infection, whereas, at
this time, the α-galactosidase
activities decreased by 70%. Interestingly, the animals died
48 h after infection, but approximately 5% of all the enzymes
tested remained active in the midgut following host death.
At this time, most of the cultivable native intestinal bacteria
had died.
[Back to top]
Albumin Competitively Inhibits Glycation of Less Abundant
Proteins
H.S. Bhonsle, S.K. Singh, G. Srivastava, R. Boppana
and M.J. Kulkarni
Glycation, a non-enzymatic reaction between glucose and
protein is the primary cause of diabetic complications. Albumin,
the most abundant plasma protein undergoes glycation both
in vivo and in vitro. The influence of albumin
on glycation of less abundant proteins has not been addressed.
For the first time, we show that albumin competitively inhibits
the glycation of less abundant proteins. This study suggests
that at least in the initial stages of diabetes, albumin may
protect other proteins from glycation.
[Back to top]
Multiple Ligands in Opioid Research
S. Ballet, M. Pietsch and A.D.
Abell
The observation in 1979 that opioid receptors interact,
led to the design of bivalent ligands in an attempt to improve
selectivity and affinity towards the different subtypes (i.e.
μ,
δ and
κ).
Dimers of monovalent “parent” opioid structures
have been evaluated and include: (a) endogenous (e.g. enkephalins)
or exogenous (e.g. dermorphin) peptide dimer analogues (b)
mixed peptidic-non-peptidic bivalent ligands and (c) dual
non-peptidic dimers. Chimeric structures, using an opioid
pharmacophore in combination with a non-opioid pharmacophore,
have also been prepared. The common aim in all these studies
is to improve the pharmacological profile of potential analgesics
to minimize common opioid-induced side-effects, such as physical
dependence and tolerance. Here we present a brief overview
of efforts to develop bivalent opioid ligands for use in pain-related
research.
[Back to top]
Opioid Peptides and Innate Immune Response in Mollusc
D.-W. Liu
The nervous and the immune systems can exchange information
through opioid peptides. Furthermore, some opioid peptides
can function as endogenous messengers of the immune system,
and participate in an important part in the regulation of
the various components of the immune response. Since the capacity
of immunocytes to release and respond to opioid neuropeptide
messengers is not restricted to mammalian organisms, recent
studies have indicated that invertebrate models have been
particularly useful to understand the mechanisms of the immune
response. Moreover, the immunocytes of molluscs resemble cells
of the vertebrate monocyte/macrophage lineage and are activated
by similar substances, which control the main immune responses,
i.e. phagocytosis, chemotaxis, and cytotoxicity. Recently,
Mytilus edulis has been the subject of recent studies
to determine whether the relationship between the immune and
nervous systems seen in vertebrates also exists in invertebrates.
The focus of this review is to describe how the opioid peptides
participate in immune processes in molluscs.
[Back to top]
Efficient Expression of Membrane-Bound Water Channel Protein
(Aquaporin Z) in Escherichia coli
J. Lian, X. Fang, J. Cai, Q. Chen, Q. Zheng,
L. Kai and Z. Xu
In order to explore the possibility of preparing a high-efficiency
aquaporin-based biofilter, an efficient approach for expression
of membrane-bound Aquaporin Z (AqpZ) in E. coli was
proposed. The AqpZ gene was amplified by means of PCR, and
two expression vectors (pET28-AqpZ and pET32-AqpZ) were constructed.
The channel protein of interest was synthesized in E.
coli BL21(DE3)/pET32-AqpZ as an insoluble fusion protein
linked with trxA. However, with BL21(DE3)/pET28-AqpZ,
significant amount of AqpZ fused only with 6-His (6-His-AqpZ)
could be expressed, correctly folded and targeted into the
membrane. Under the optimized culture conditions, the highest
expression level (9.05 mg/l) of membrane-bound 6-His-AqpZ
was achieved with BL21(DE3)/pET28-AqpZ, and an additional
amount (2.35 mg/l) was expressed concomitantly as the inclusion
body form. This expression result was 3.5 times higher than
that in the previous studies.
[Back to top]
A Protein Interaction Network Analysis for Yeast Integral
Membrane Protein
M.-G. Shi, D.-S. Huang and X.-L. Li
Although the yeast Saccharomyces cerevisiae
is the best exemplified single-celled eukaryote, the vast
number of protein-protein interactions of integral membrane
proteins of Saccharomyces cerevisiae have not been
characterized by experiments. Here, based on the kernel method
of Greedy Kernel Principal Component analysis plus Linear
Discriminant Analysis, we identify 300 protein-protein interactions
involving 189 membrane proteins and get the outcome of a highly
connected protein-protein interactions network. Furthermore,
we study the global topological features of integral membrane
proteins network of Saccharomyces cerevisiae. These
results give the comprehensive description of protein-protein
interactions of integral membrane proteins and reveal global
topological and robustness of the interactome network at a
system level. This work represents an important step towards
a comprehensive understanding of yeast protein interactions.
[Back to top]
A New Family of Small (4kDa) Neurotoxins from the Venoms of
Spiders of the Genus Phoneutria
A.D. Lúcio, F.V. Campos, M. Richardson,
M.N. Cordeiro, M.S.C. Mazzoni, M.E. de Lima, A.M.C. Pimenta,
M.P. Bemquerer, S.G. Figueiredo, P.C. Gomes and P.S.L.
Beirão
A family of 4kDa neurotoxic peptides was purified from
venoms of Phoneutria spiders. All have six cysteine
residues, and low similarity with other neurotoxins. Three
toxins caused moderate inhibition of L-type Ca2+
channels. The structure of toxin PRTx27C3 was modeled and
compared with toxin ADO1. The importance of four residues
is suggested.
[Back to top]
Crystal Structure of SCO6571 from Streptomyces coelicolor
A3(2)
P. Begum, N. Sakai, T. Hayashi, Y.-G. Gao,
T. Tamura, N. Watanabe, M. Yao and I. Tanaka
SCO6571 protein from Streptomyces coelicolor
A3(2) was overexpressed and purified using Rhodococcus
erythropolis as an expressing host. Crystals of selenomethionine-substituted
SCO6571 have been obtained by vapor diffusion method. SCO6571
crystals diffract to 2.3 Å
and were found to belong to the orthorhombic space group P212121
with unit cell parameters α
= 84.5, b = 171.6, c = 184.8 Å.
Six molecules in the asymmetric unit give a crystal volume
per protein mass (VM)
of 2.97 Å3
Da-1 and solvent content
of 58.6 %. The structure was solved by the single wavelength
anomalous diffraction (SAD) method. SCO6571 is a TIM-barrel
fold protein that assembles into a hexameric molecule with
D3 symmetry.
[Back to top]
Structural and Spectroscopic Elucidation of Tetrapetide Glycyl
-L-Prolyl-Glycyl-Glycine
and Its Hydrogensquarate
T.M. Kolev
The IR-spectroscopic and structural elucidation of tetrapeptide
glycyl-L-prolyl-glycyl-glycine
and its hydrogensquarate was performed by employing linear-polarized
IR-spectroscopy of oriented colloid suspensions in nematic
host as well as mass spectrometry. Quantum-chemical ab
initio calculations were carried out in order to evaluate
both the electronic structure and optical properties of the
compound studied.
[Back to top]
Yellow Lupine Cyclophilin Interacts with Nucleic Acids
K. Nuc, K. Lesniewicz, P. Nuc and
R. Slomski
To investigate properties of yellow lupine cytosolic
cyclophilin, an expression vector pET15CYP was constructed.
The CyP cDNA (GenBank accession no.Y16088) reveals an open
reading frame of 172 amino acids with the conserved tryptophan
residue at position 128 and an insertion of seven amino acids
spanning positions 48-54. Yellow lupine cyclophilin, purified
after expression in E. coli cells, exhibits peptidyl-prolyl
cis/trans isomerase activity when assayed with a
synthetic oligopeptide. We have demonstrated that the recombinant
cyclophilin is able to interact with nucleic acids, both single
and double stranded DNA fragments as well as RNA.
[Back to top]
Purification, Biochemical and Functional Characterization
of Miliin, a New Thiol-Dependent Serine Protease Isolated
from the Latex of Euphorbia milii
L.P. Moro, M.T. Murakami, H. Cabral, A.
Vidotto, E.H. Tajara, R.K. Arni, L. Juliano and G.O.
Bonilla-Rodriguez
Miliin, a new thiol-dependent serine protease purified
from the latex of Euphorbia milii possesses a molecular
weight of 79 kDa, an isoelectric point of 4.3 and is optimally
active at 60 ºC in the pH range of and 7.5-11.0. Activity
tests indicate that milliin is a thiol-dependent serine protease.
[Back to top]
Three Sampling Strategies to Predict Mutations in H5N1 Hemagglutitins
from Influenza A Virus
G. Wu and S. Yan
After several studies on prediction of mutation, we examine
the effect of three sampling strategies, the sampling based
on years, the sampling based on number of mutations, and the
sampling based on the unpredictable portion of amino-acid
pairs, on the prediction performance in H5N1 hemagglutinins.
The results show that the sampling strategy does play an important
role in prediction, which should be taken into account when
predicting the next generation of mutations in proteins from
influenza A virus.
[Back to top]
Predicting Subcellular Localization of Mycobacterial Proteins
by Using Chou’s Pseudo Amino Acid Composition
H. Lin, H. Ding, F.-B. Guo, A.-Y. Zhang
and J. Huang
The successful prediction of protein subcellular localization
directly from protein primary sequence is useful to protein
function prediction and drug discovery. In this paper, by
using the concept of pseudo amino acid composition (PseAAC),
the mycobacterial proteins are studied and predicted by support
vector machine (SVM) and increment of diversity combined with
modified Mahalanobis Discriminant (IDQD). The results of jackknife
cross-validation for 450 non-redundant proteins show that
the overall predicted successful rates of SVM and IDQD are
82.2% and 79.1%, respectively. Compared with other existing
methods, SVM combined with PseAAC display higher accuracies.
[Back to top]
Construction, Expression, Purification and Immunology Effect
of an Anti-Atherosclerosis Chimeric Enzyme Vaccine in Escherichia
coli
S. Yunxiao, L. Zhufang, Y. Xin, L. Jun,
X. Qiyan, Q. Gaofu, C. Rongyue, W. Jie, L. Taiming, F. Hao
and L. Jingjing
In order to prevent atherosclerosis, a chimeric enzyme vaccine
of AnsB-TTP-PADRE-CETPC was successfully constructed, expressed
and purified to immunize New Zealand white rabbits for inducing
high titers of anti-CETP antibodies to improve lipid abnormality.
The protein was expressed as soluble protein in Escherichia
coli and purified by anion exchange column and Sephadex
G-100 size-exclusion chromatography. After immunizing rabbits
with the purified protein, high titer anti-CETP antibodies
were induced and lasted more than nineteen weeks in vivo;
High density lipoprotein cholesterol (HDL-C) content in the
serum was elevated to 61% while decreased low density lipoprotein
cholesterol (LDL-C) to 37.2% compared with control rabbits
in the presence of Al(OH)3.
[Back to top]
Preliminary Structural Studies on MPN423 Expressed from an
Orthologous ORFan of Mycoplasma pneumoniae
D.H. Shin
ORFans are orphan open reading frames. The numbers of
ORFans are steadfastly increasing despite of the genome database
increment. Characterizing ORFans is essential to fully understanding
the diversity of the structure and function of proteins in
nature. In this study, MPN423 from Mycoplasma pneumoniae
has been cloned, expressed, purified, and crystallized. MPN423
is an orthologous ORFan whose only known homologue in the
whole genome database is MG296 from M. genitalium.
X-ray diffraction data were collected to 2.7 Å
from the crystal of a selenomethionine substitute MPN423.
The crystal belongs to the primitive monoclinic space group
P21, with unit-cell parameters
of a = 50.5 Å,
b = 89.2 Å,
c = 50.6 Å,
and β
= 102.9
γ. Å
preliminary electron density map shows five α-helical
segments per MPN423 molecule. A full structure determination
is under way to provide helpful information to general questions
about orthologous ORFan products.
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