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Protein
& Peptide Letters
ISSN: 0929-8665
Protein & Peptide Letters
Volume 15, Number 3, 2008
Contents
Regular Papers
Host Defense Peptides and the New Line of Defence
Against Multiresistant Infections Pp. 238-243
T. Hirsch, F. Jacobsen, H.-U. Steinau and L. Steinstraesser
[Abstract]
Conformational Studies Suggest That the Double
Stranded β
Helix Scaffold Provides an Optimal Balance Between Protein
Stability and Function Pp. 244-249
M. Rajavel, N.N. Kulkarni and B. Gopal
[Abstract]
Normal Cytochrome c Oxidase Activity in Prion
Protein Gene-Deficient Mice Pp. 250-254
A. Sakudo, Y. Taniuchi, T. Kobayashi, T. Onodera and K.
Ikuta
[Abstract]
Stability and Biological Activity of Human Intestinal
Trefoil Factor Produced by Pichia pastoris
Pp. 255-259
Y. Sun, X. Peng, Y. Zhang, S. Lv, W. Wu and S. Wang
[Abstract]
Designing of Substrates and Inhibitors of Bovine α-Chymotrypsin
with Synthetic Phenylalanine Analogues in Position P1
Pp. 260-264
M. Wysocka, A. Lesner, A. Legowska, A. Jaskiewicz, H.
Miecznikowska and K. Rolka
[Abstract]
Surfactin Isoforms from Bacillus subtilis
HSO121: Separation and Characterization Pp. 265-269
N.I.A. Haddad, X. Liu, S. Yang and B. Mu
[Abstract]
Effects of a Chitin Binding Vicilin from Erythrina
velutina Seeds on Bean Bruchid Pests (Callosobruchus
maculatus and Zabrotes subfasciatus) Pp.
270-274
F.M. Teixeira, A.S. Oliveira, L.L.P. Macedo, E.A. Santos
and M.P. de Sales
[Abstract]
Structure-Activity Relationships of the Dimeric
Analogues of Endomorphin-2 with Different Lengths of Spacers
Pp. 275-279
Y.-F. Gao, M.-X. Zhai, W.-X. Liu, X. Liu, Y. Yuan, Y.-M.
Qi and R. Wang
[Abstract]
T and B-Cell Epitopes Prediction of Iranian Saffron
(Crocus sativus) Profilin by Bioinformatics Tools
Pp. 280-285
B. Saffari, H. Mohabatkar and S. Mohsenzadeh
[Abstract]
Predicting Subcellular Localization with AdaBoost
Learner Pp. 286-289
Y.-h. Jin, B. Niu, K.-Y. Feng, W.-C. Lu, Y.-D. Cai and
G.-Z. Li
[Abstract]
A Docking Modelling Rationally Predicts Strong Binding
of Bisphenol A to Estrogen-Related Receptor γ
Pp. 290-296
T. Nose and Y. Shimohigashi
[Abstract]
Stabilization of Beta-Propeller Phytase by Introducing
Xaa→Pro
and Gly→Ala
Substitutions at Consensus Positions Pp. 297-299
E.T.K. Tung, H.-W. Ma, C. Cheng, B.L. Lim and K.-B. Wong
[Abstract]
Recent Advances and Perceptions in Studies of
Heterodimerization Between G Protein-Coupled Receptors
Pp. 300-308
H. Satake and T. Sakai
[Abstract]
Linear-Polarized IR-Spectroscopic and Structural
Elucidation of Glycyl-L-Phenylalanyl-Glycine
Hydrochloride Monohydrate Pp. 309-313
B.B. Koleva
[Abstract]
Diagnostic Utility of Anti-Cyclic Citrullinated Peptide
and Anti-Modified Citrullinated Vimentin Antibodies in Rheumatoid
Arthritis Pp. 314-317
G. Keskin, A. Inal, D. Keskin, A. Pekel, O. Baysal, U.
Dizer and A. Sengül
[Abstract]
Crystallization Report
Purification, Crystallization and Preliminary Analysis
of Hemoglobin from Rabbit (Oryctolagus cuniculus)
Pp. 318-319
S. Sundaresan, P. Charles, K. Neelagandan and M.N.G. Ponnuswamy
[Abstract]
Abstracts
[Back to top]
Host Defense Peptides and the New Line of Defence
Against Multiresistant Infections
T. Hirsch, F. Jacobsen, H.-U. Steinau and L. Steinstraesser
Increasing antibiotic resistance has led to an urgent
need for new therapeutic approaches. Host defense peptides
are known to be antimicrobial and have revealed broad immunomodulatory
functions for both innate and adaptive immunity. This review
will focus on the role of host defense peptides in infection
and immune response and discuss its potential and limitations
as a future therapeutical agent.
[Back to top]
Conformational Studies Suggest That the Double Stranded β
Helix Scaffold Provides an Optimal Balance Between Protein
Stability and Function
M. Rajavel, N.N. Kulkarni and B. Gopal
Proteins with the double stranded beta-helix (DSBH, also
known as cupin) fold perform a diverse range of functions.
In this study, Bacillus subtilis quercetinase was
used as a model system to understand the conformational determinants
of functional diversity within the cupin fold. Controlled
proteolysis experiments revealed that this enzyme is active,
thermo-stable and maintains its quaternary arrangement even
after substantial (ca 33 %) cleavage of the protein. The results
presented in this manuscript thus show that the DSBH scaffold
offers a novel balance between protein stability and function
by locating the active site and substrate recognition features
in the most stable region of the protein.
[Back to top]
Normal Cytochrome c Oxidase Activity in Prion Protein Gene-Deficient
Mice
A. Sakudo, Y. Taniuchi, T. Kobayashi, T. Onodera and K.
Ikuta
Cytochrome c oxidases of prion protein (PrP) gene-deficient
(Prnp -/-)
and Prnp +/+
mice were examined in vivo and in vitro.
Non-invasive near-infrared spectra revealed that oxidation
of copper and heme a+a3
in cytochrome c oxidase of Prnp -/-
mice was similar to that in Prnp +/+
mice. Biochemical analysis of mitochondrial fractions also
supported the results. PrP might not be involved in regulation
of cytochrome c oxidase.
[Back to top]
Stability and Biological Activity of Human Intestinal Trefoil
Factor Produced by Pichia pastoris
Y. Sun, X. Peng, Y. Zhang, S. Lv, W. Wu and S. Wang
Intestinal trefoil factor (ITF), a member of the trefoil
factor family, plays an important role in protecting the epithelial
layer of the intestinal tract from damage and repairing epithelium
after injury. In the present study, we expressed recombinant
hITF by Picha pastoris expression system in shake
flasks to 50mg/L. rhITF was purified up to 95% by ammonium
sulfate precipitation, Ni–NTA affinity chromatography
and ultrafiltration. In conditions simulating intestinal tract
environment, rhITF was shown to exhibit resistance to proteases,
heat, acid and alkali. In addition, rhITF was found to be
biologically active in an in vitro restitution model.
This study lays the foundation for the commercial production
of rhITF and provides theoretical evidence for the oral use
of rhITF.
[Back to top]
Designing of Substrates and Inhibitors of Bovine α-Chymotrypsin
with Synthetic Phenylalanine Analogues in Position P1
M. Wysocka, A. Lesner, A. Legowska, A. Jaskiewicz, H.
Miecznikowska and K. Rolka
The primary specificity residue of a substrate or an
inhibitor, called the P1
residue, is responsible for the proper recognition by the
cognate enzyme. This residue enters the S1
pocket of the enzyme and establishes contacts
(up to 50%) inside the proteinase substrate cavity, strongly
affecting its specificity. To analyze the influence on bovine
α -chymotrypsin
substrate activity, aromatic non-proteinogenic amino acid
residues in position P1 with
the sequence Ac-Phe-Ala-Thr-X-Anb5,2
-NH2 were introduced:
L-pyridyl alanine (Pal), 4-nitrophenylalanine – Phe(p-NO2),
4-aminophenylalanine – Phe(p-NH2),
4-carboxyphenylalanine Phe(p-COOH), 4-guanidine phenylalanine
– Phe(p-guanidine), 4-methyloxycarbonyl-phenylalanine
– Phe(p-COOMe), 4-cyanophenylalanine –
Phe(p-CN), Phe, Tyr. The effect of the additional
substituent at the phenyl ring of the Phe residue was investigated.
All peptides contained an amide of 5-amino-2-nitrobenzoic
acid, which served as a chromophore. Kinetic parameters (kCat
KM and kCatt/KM)
of the peptides synthesized with bovine α
-chymotrypsin were determined. The highest value of
the specificity constant kCat/KM,
reaching 6.0×105 [M-1
×s -1],
was obtained for Ac-Phe-Ala-Thr-Phe(p-NO2)-Anb5,
2 -NH2. The
replacement of the acetyl group with benzyloxycarbonyl moiety
yielded a substrate with the value of kCat
more than three times higher. Peptide aldehydes were synthesized
with selected residues (Phe, Pal, Tyr, Phe(p-NO2)
in position P1 and potent
chymotrypsin inhibitors were obtained. The dissociation constant
(K i) with the experimental
enzyme determined for the most active peptide, Tos-Phe-Ala-Thr-Phe(p-NO2)-CHO,
amounted to 1.12×10-8
M.
[Back to top]
Surfactin Isoforms from Bacillus subtilis HSO121:
Separation and Characterization
N.I.A. Haddad, X. Liu, S. Yang and B. Mu
Natural surfactin is a mixture of cyclic lipopeptides
built from variants of a heptapeptide and a β-hydroxy
fatty acid. A biosurfactant-producing strain, Bacillus
Sabtilis HSO121, was isolated from the production water
of an oil field. The strain was able to produce eight surfactin
isoforms which have been isolated by acid-precipitation followed
by extraction into methanol. A novel procedure for the purification
of surfactin was achieved. It consists of a solid-phase extraction
on C18 gel followed by reversed-phase
high performance liquid chromatography using Prep. HiQ sil
C18W, column. The surfactin isoforms were eluted by linear
acetonitrile gradient from 80-100%. The peaks were analyzed
by TLC on silica gel, and after acid hydrolysis their amino
acid compositions were determined by HPLC analysis. Eight
isoforms of surfactin had nearly the same amino acid composition
and appeared a single spot in TLC. According to the Rf
values with the amino acid composition, these peaks belong
to the surfactin group of lipopeptides. Infrared analysis
of the purified samples also revealed a pattern similar to
that of surfactin. This is a very effective method for isolating
and fractionating lipopeptides, of the same or different nature.
[Back to top]
Effects of a Chitin Binding Vicilin from Erythrina velutina
Seeds on Bean Bruchid Pests (Callosobruchus maculatus
and Zabrotes subfasciatus)
F.M. Teixeira, A.S. Oliveira, L.L.P. Macedo, E.A. Santos
and M.P. de Sales
Erythrina velutina vicilin, EvV, is a dimeric glycoprotein
with Mr of 124.6 kDa. EvV was tested for anti-insect activity
against bean bruchid larvae. EvV had LD50
of 0.10% and ED50
of 0.14% for Z. subfasciatus and LD50
of 0.26% and ED50 of 0.19%
for C. maculatus. EvV was not digested by bean larvae
enzymes until 12 h of incubation, and at 24 h EvV was more
resistant to Z. subfasciatus enzymes.
[Back to top]
Structure-Activity Relationships of the Dimeric Analogues
of Endomorphin-2 with Different Lengths of Spacers
Y.-F. Gao, M.-X. Zhai, W.-X. Liu, X. Liu, Y. Yuan, Y.-M.
Qi an R. Wang
In the present study, seven novel dimeric analogues of
endomorphin-2 with longer spacers were designed and synthesized.
Through dimerization, their affinity for δ-opioid
receptor was mostly increased, especially the δ-opioid
receptor preferred dimeric analogue, DEM12.
The results were confirmed by the in vitro bioassay.
The structure-activity relationships were also discussed.
[Back to top]
T and B-Cell Epitopes Prediction of Iranian Saffro (Crocus
sativus) Profilin by Bioinformatics Tools
B. Saffari, H. Mohabatkar and S. Mohsenzadeh
Plant profilins form a well-known panallergen family
responsible for cross-sensitization between plant foods and
pollens. We sought to map T and B-cell epitopes on the Iranian
Crocus sativus profilin by bioinformatics tools.
The predicted peptides are useful for further vaccine development.
[Back to top]
Predicting Subcellular Localization with AdaBoost Learner
Y.-h. Jin, B. Niu, K.-Y. Feng, W.-C. Lu, Y.-D. Cai and
G.-Z. Li
Protein subcellular localization, which tells where a
protein resides in a cell, is an important characteristic
of a protein, and relates closely to the function of proteins.
The prediction of their subcellular localization plays an
important role in the prediction of protein function, genome
annotation and drug design. Therefore, it is an important
and challenging role to predict subcellular localization using
bio-informatics approach. In this paper, a robust predictor,
AdaBoost Learner is introduced to predict protein subcellular
localization based on its amino acid composition. Jackknife
cross-validation and independent dataset test were used to
demonstrate that Adaboost is a robust and efficient model
in predicting protein subcellular localization. As a result,
the correct prediction rates were 74.98% and 80.12% for the
Jackknife test and independent dataset test respectively,
which are higher than using other existing predictors. An
online server for predicting subcellular localization of proteins
based on AdaBoost classifier was available on http://chemdata.shu.
edu.cn/sl12.
[Back to top]
A Docking Modelling Rationally Predicts Strong Binding of
Bisphenol A to Estrogen-Related Receptor γ
T. Nose and Y. Shimohigashi
A computer-aided docking study was carried out to quickly
clarify the binding structure of the ligand-receptor complex
between bisphenol A (BPA), a well-known endocrine disruptor,
and estrogen-related receptor γ
(ERRγ).
The resulting complex indicated that BPA binds to the ligand-binding
pocket of ERR γ
without any disruptions of the activation conformation.
[Back to top]
Stabilization of Beta-Propeller Phytase by Introducing Xaa→Pro
and Gly→Ala
Substitutions at Consensu Positions
E.T.K. Tung, H.-W. Ma, C. Cheng, B.L. Lim and K.-B. Wong
Phytases play important roles in agricultural and feed
industries. In this study, the stability of a beta-propeller
phytase, PhyL, from Bacillus licheniformis was successfully
improved by introducing Xaa→Pro
and Glyà→Ala
substitutions at consensus positions. Our results suggest
that Glyà→Ala
substitution is a more promising strategy to improve protein
stability.
[Back to top]
Recent Advances and Perceptions in Studies of Heterodimerization
Between G Protein-Coupled Receptors
H. Satake and T. Sakai
G pretein coupled-receptors (GPCR) have been shown to
form heterodimers or heterooligomers with various biochemical
and/or pharmacological activity that are distinct from those
of the corresponding monomers or homomers. Recent in vitro
and in vivo experimental data have also shown novel
functional roles of several orphan GPCRs as modulatory units
of heterodimers and potentials for development of heterodimer-selective
clinical agents. In this review, we summarize essential and
latest knowledge as to GPCR heterodimerizations and the current
issues to be solved in the near future.
[Back to top]
Linear-Polarized IR-Spectroscopic and Structural Elucidation
of Glycyl-L-Phenylalanyl-Glycine
Hydrochloride Monohydrate
B.B. Koleva
As a part of our spectroscopic and structural elucidation
of small peptides, their salts and metal complexes, the polarized
IR-spectroscopic and structural elucidation of the protonated
form of glycyl- L -phenylalanyl-glycine
as hydrochloride monohydrate (H-Gly-Phe-Gly-OHxHClxH2O)
is reported. The obtained structure of the protonated tripeptide
is supported by quantum chemical ab initio calculations at
UHF level of theory and 6-31++G**, showing only one stable
conformer with two intermolecular NH3+…OH2
and (C=O)OH…OH2 hydrogen
bonds. The Nterminus amide
O=C-N-H amide fragment is flat trans-configured with
a dihedral angle value of 179.0(7)o,
while the corresponding group at Cterminus
is with transoide-configurated and value of the dihedral
angle of 151.1(3)o, respectively.
In addition data of 1H- and
13 C magnetic resonance spectroscopy
(NMR), thermogravimetry (TGA), differential scanning calorimetry
(DSC) and high performed liquid chromatography (HPLC) with
tandem mass spectrometry (HPLC-MS/MS) as presented.
[Back to top]
Diagnostic Utility of Anti-Cyclic Citrullinated Peptide and
Anti-Modified Citrullinated Vimentin Antibodies in Rheumatoid
Arthritis
G. Keskin, A. Inal, D. Keskin, A. Pekel, O. Baysal, U.
Dizer and A. Sengül
Several autoantibodies found in RA are directed to epitopes
in citrullinated proteins. One of them is anti modified citrullinated
vimentin (Anti-MCV). We tested the value a newly developed
ELISA for the detection of antibodies againts a genetically
modified citrullinated vimentin (anti-MCV) in comparison with
an anti-CCP based ELISA system for the diagnosis of RA.
Thirty-five patients with RA (mean age; 42.6 ± 10.87
years, mean disease duration; 9.37 ± 3.98 years) were
enrolled in this study. Twenty –five ankylosing spondylitis
(mean age; 35.88 ± 6.64 years, mean disease duration;
10.25 ± 4.61 years), and 19 healthy subjects (mean
age; 40.26 ± 5.11 years) served as controls. Anti-CCP
antibodies and Anti-MCV antibodies were measured using ELISA.
In all RA patients, mean anti- CCP level was 69.07 ±
90.43 U/ml and anti-MCV level was 665.77 ± 1040.19
U/ml. In pa-tients with AS, the mean anti-CCP level was 10.7
± 5.22 U/ml and anti-MCV level was 40.54 ± 20.15
U/ml. In healthy controls, the mean anti-CCP level was 11.11
± 7.65 U/ml, anti-MCV level was 23.12 ± 12.04
U/ml. In patients with active RA, the mean serum anti-CCP
level was 100.54 ± 98.07 U/ml and anti-MCV level was
998.74 ± 1154.93 U/ml. In patients with inactive RA,
the mean serum anti-CCP level was 8.77 ± 1.55 U/ml
and anti-MCV level was 27.59 ± 23.10 U/ml.
According to these results; In patients with RA, the mean
serum anti-MCV and anti-CCP levels were significantly high
compared to patients with AS and healthy controls (p=0.002,
p=0.001, p=0.002, p=0.001 respectively). The mean serum anti-MCV
and anti- CCP levels were significantly higher in active patients
with RA than in inactive patients with RA patients (p=0.001
and p=0.001 respectively). In inactive patients with RA, the
mean serum anti-MCV and anti-CCP levels were similar in patients
with AS and patients (p=0.484, p=0.308, p=0.09 and p=0.222
respectively).
The mean serum anti-MCV levels were correlated with DAS 28
(r=0.531, p=0.001), VAS score (r=0.332, p=0.01), ESR (r=0.458,
p=0.001), serum CRP levels (r=0.568, p=0.01), serum RF levels
(r=0.529, p=0.001), swollen joints number (r=0.525, p=0.001)
and tender joints number (r=0.638, p=0.001).
As a result; measurement of serum anti-MCV levels is useful
for diagnosis of RA and combined use of anti-MCV and RF may
be more useful prognostic factor than either method alone,
RF and anti-CCP.
[Back to top]
Purification, Crystallization and Preliminary Analysis of
Hemoglobin from Rabbit (Oryctolagus cuniculus)
S. Sundaresan, P. Charles, K. Neelagandan and M.N.G. Ponnuswamy
Hemoglobin (Hb) is a tetrameric protein, which contains
four heme prosthetic groups, and each one is associated with
a polypeptide chain. Herein, we report the rabbit hemoglobin
which has intrinsically high oxygen affinity and possess highest
sequence identity with human hemoglobin. The purified hemoglobin
has been tried to crystallize in different crystallization
conditions owing to its formation of various crystal systems.
The rabbit Hb crystals were grown using PEG 3350 as the precipitant
at 18oC. The crystals of
rabbit Hb belongs to triclinic space group P1 with one molecule
(α2β2)
in the asymmetric unit.
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