Protein & Peptide Letters, Volume 9, No. 5, 2002
Protein Structure to Function via Dynamics Pp.367-377
Neeti
Sinha , and Sandra J. Smith-Gill
The use of DODT as a non-malodorous scavenger
in Fmoc-based peptide synthesis Pp.379-385
A.
Teixeira, W.E. Benckhuijsen, P.E. de Koning, A.R.P.M. Valentijn and J.W.
Drijfhout
Kinetic Analysis of the Cleavage of Human
Protease-Activated Receptor-1 / 2 / 3 and 4 Using Quenched-Fluorescent Peptide Substrates Pp.387-393
Mark
T. Fox , Brett Greer, Jill Lawson, Adrienne Healy and Patrick Harriott.
Antibiotic Activity of Reversed Peptides of a-Helical Antimicrobial Peptide, P18 Pp.395-402
Si-Hyung
Lee , Dong Gun Lee , Sung-Tae Yang , Yangmee Kim ,Jae Il Kim , Kyung-Soo
Hahm & Song Yub Shin
Conformational Analysis of Sea Cucumber
(Caudina arenicola) C Globin 94-97 Fragment , Pro-Glu-Leu-Leu Pp.403-409
Sunil
Kumar, P. N., Devaky, K. S., 1Sadasivan, C. and Haridas, M.
A Synthetic Strategy for ON-Resin Amino
Acidspecific Multiple Fatty Acid Acylation of Peptides Pp.411-417
Avaronnan
Harishchandran, Bhattaram Pallavi, Ramakrishnan Nagaraj
Cloning and High-Level Expression of Scorpion
Toxin BmKITa1 in Escherichia coli and Insect Cells Pp.419-426
Zhen
Liu, Guan-Zhen Yang, Cheng-Wu Chi, Xiang-Fu Wu
Functionally Important Residues for the
Anticoagulant Activity of a Basic Phospholipase A2 from the Agkistrodon halys
pallas Pp.427-434
Xiaoyan
Zhong, Haomang Jiao, Liang Fan, Xiangfu
Wu and Yuancong Zhou
Effects of Stereochemistry of Sugars on
Protein Stabilities Pp.435-439
Kiyonari
Masumoto, Tadashi Ueda , Makiko Nagata, Yuko Yamada,Yuichiro Yoshida, Yoshio
Hashimoto, and Taiji Imoto
Purification, Crystallization and Initial
Structural Solution of a New a-Like Toxin with Cardiac Toxicity from
Scorpion Buthus martensii Karsch Pp.441-449
Rong-Jin
Guan, Xiao-Lin He, Miao Wang, Ye-Xiang, Gen-Pei Li and Da-Cheng Wang
Binding Potency of Peptide Fragments of Type
1 Collagen Cross-Linked N-Telopeptide Measured by an Enzyme-Linked Immunosorbant
Assay Pp.451-457
Feng-Di
T. Lung , Chiu-Heng Chen1, Heuy-Yi Chen , Chien-Chung Liou , and Yen-Meng Liou
Crystallization and preliminary X-ray
diffraction analysis of FKBP12 complexed with a novel neurotrophic ligand Pp.459-463
Pengyun Li , Liwei Wang , Yi Ding , Beili Wu , Cuiling Shu , Aihua Nie , Song Li , Beifen Shen & Zihe Rao
[Back to top] Protein Structure to Function via Dynamics
Neeti
Sinha , and Sandra J. Smith-Gill
Protein folding,
binding, catalytic activity and molecular recognition all involve molecular
movements, with varying extents. The molecular movements are brought upon via
flexible regions. Stemming from sequence, a fine tuning of electrostatic and
hydrophobic properties of the protein fold determine flexible and rigid
regions. Studies show flexible regions usually lack electrostatic interactions,
such as salt-bridges and hydrogen-bonds, while the rigid regions often have
larger number of such electrostatic interactions. Protein flexible regions are
not simply an outcome of looser packing or instability, rather they are
evolutionally selected. In this review article we highlight the significance of
protein flexibilities in folding, binding and function, and their structural
and thermodynamic determinants. Our electrostatic calculations and molecular
dynamic simulations on an antibody-antigen complex further illustrate the
importance of protein flexibilities in binding and function.
[Back to top] The use of DODT as a non-malodorous scavenger
in Fmoc-based peptide synthesis
A. Teixeira, W.E. Benckhuijsen, P.E. de Koning, A.R.P.M. Valentijn and J.W. Drijfhout
We have
synthesized a random group of peptides and performed cleavages using various
cleavage cocktails including 3,6-dioxa-1,8-octanedithiol (DODT). Purity of the
peptides was compared to that obtained with standard protocols for cleavage
using RP-HPLC and Maldi-Tof mass spectrometry. We show that stinking thiols
can be replaced by the almost odourless
(DODT) without negatively affecting the purity of the end product.
[Back to top] Kinetic Analysis of the Cleavage of Human
Protease-Activated Receptor-1 / 2 / 3 and 4 Using Quenched-Fluorescent Peptide
Substrates
Mark
T. Fox , Brett Greer, Jill Lawson, Adrienne Healy and Patrick Harriott.
Protease-activated
receptors [PARs] are a family of G-protein-coupled seven-transmembrane domain
receptors that are activated by proteolytic cleavage of their amino-terminal
exodomain. To characterize the cleavage rate of human PAR-1 / 2 / 3 and 4 by
trypsin and thrombin, four synthetic quenched-fluorescent peptide substrates
have been synthesized. Each substrate consisted of a ten-residue peptide
spanning the receptor activation cleavage site and using progress-curve
kinetics, kcat / Km values were determined.
[Back to top]
Antibiotic Activity of Reversed Peptides of a-Helical Antimicrobial Peptide, P18
Si-Hyung
Lee , Dong Gun Lee , Sung-Tae Yang , Yangmee Kim ,Jae Il Kim , Kyung-Soo
Hahm & Song Yub Shin
P18
(KWKLFKKIPKFLHLAKKF-NH2), an a-helical antimicrobial peptide designed from cecropin Amagainin 2 hybrid, was known to have potent antimicrobial activity
against bacteria as well as fungi without hemolytic activity. To find the
peptides comparable or superior to the antimicrobial activity of P18, the two
reversed peptides (Rev-1 and Rev-2) of P18 were designed and synthesized. These
peptides were found to have similar antimicrobial activity against bacterial
and fungal cells without hemolytic activity as compared with P18. Furthermore,
a reversed peptide, Rev-2 was shown to have a two-fold higher activity in
killing some bacterial cells than P18. Therefore, these results suggested that
Rev-2 peptide seems to be an excellent candidate for developing novel peptide
antibiotics.
[Back to top] Conformational Analysis of Sea Cucumber
(Caudina arenicola) C Globin 94-97 Fragment , Pro-Glu-Leu-Leu
Sunil
Kumar, P. N., Devaky, K. S., 1Sadasivan, C. and 1Haridas, M.
The tetrapeptide
Pro-Glu-Leu-Leu forms the 94-97 fragment of C globin in sea cucumber. 2%
Butanediol dimethacrylate-cross linked polystyrene (2% BDDMA-PS), which had
been optimized, was used for the synthesis of the tetrapeptide Pro-Glu-Leu-Leu.
The peptide was synthesized by using Boc-amino acid strategy. The peptide
purity was checked by RP-HPLC and the peptide was characterized by 1H NMR
spectroscopy and amino acid analysis. Conformation of the peptide was studied
by 1D- and 2D- homonuclear 1H NMR, in DMSO-d6 at 300K. The conformation of the
synthetic tetrapeptide (extended backbone conformation) is not in agreement
with that in C globin.
[Back to top] A Synthetic Strategy for ON-Resin Amino
Acidspecific Multiple Fatty Acid Acylation of Peptides
Avaronnan
Harishchandran1, Bhattaram Pallavi1, Ramakrishnan Nagaraj
Covalent
modification with fatty acids is observed in several proteins that play crucial
roles in cellular physiology. In this paper, a convenient method for the
generation of multiple fatty acylated synthetic peptides is described. Peptides
were synthesized using solid phase procedures with fluorenylmethoxycarbonyl
a-amino protected amino acids. Acetamidomethyl protected cysteines were employed.
The thiol protecting group was selectively deprotected and acylation was
carried out on the resin-bound peptides. The strategy described in this report
is applicable to any peptide sequence.
[Back to top] Cloning and High-Level Expression of Scorpion
Toxin BmKITa1 in Escherichia coli and Insect Cells
Zhen
Liu, Guan-Zhen Yang, Cheng-Wu Chi, Xiang-Fu Wu
BmK ITa1 cDNA was
cloned and highly expressed in E. coli and insect cell. SDS-PAGE and western
blot analysis revealed that subunit molecular weight of expression products is
about 40 kDa and 10 kDa respectively. The expression product purified by a Ni2+-IDA-sepharose 6B column was toxic for insect, which indicated that it was
biologically activity. Furthermore, Quantitative estimation show that the
biological activity of recombinant BmK ITa1 from Tn cells was more powerful
than from E. coli.
[Back to top] Functionally Important Residues for the
Anticoagulant Activity of a Basic Phospholipase A2 from the Agkistrodon halys
pallas
Xiaoyan
Zhong, Haomang Jiao, Liang Fan, Xiangfu
Wu and Yuancong Zhou
To identify the
anticoagulant region of the phospholipase A2 (PLA2) from the Agkistrodon halys
Pallas (class II), four mutants E53G, W70M, T56K, and D67K were produced
according to the prediction from the crystal structure and the sequence
comparison of the strong, weak and non-anticoagulant PLA2s. A test of blood
clotting revealed that E53G and W70M had lost their effects on the blood
clotting, while T56K and D67K had enhanced activity. The four residues are
located on the same face in the tertiary structure of this enzyme. The result
supported the prediction that there exists an anticoagulant region that is
composed of some residues that are close to each other in tertiary structure to
form a functional face.
[Back to top] Effects of Stereochemistry of Sugars on
Protein Stabilities
Kiyonari
Masumoto, Tadashi Ueda , Makiko Nagata, Yuko Yamada,Yuichiro Yoshida, Yoshio
Hashimoto, and Taiji Imoto
We investigated
thermal stabilities of four proteins in the presence of four kinds of sugars to
analyze the mechanism of stabilization of proteins by additives. These proteins
were stabilized by the addition of sugars, and the degree of stabilization
correlated to the partial molar isentropic compressibility of the sugar.
[Back to top] Purification, Crystallization and Initial
Structural Solution of a New a-Like Toxin with Cardiac Toxicity from Scorpion
Buthus martensii Karsch
Rong-Jin
Guan, Xiao-Lin He, Miao Wang, Ye-Xiang, Gen-Pei Li and Da-Cheng Wang
An
a-like toxin
named BmK M7 active on both mammals and insects has been purified from the
venom of scorpion Buthus martensii Karsch (BmK) recently. The
electrophysiological experiments showed that M7 can bind to human cardiac
Na+-channel and modify its normal properties, hence can be considered as a cardiotoxin. Single crystals of M7 have been obtained by hanging-drop vapor
diffusion method using ammonium sulfate as precipitant in Tris-HCl buffer at pH
8.5. A data set to 1.40 Å resolution was collected using synchrotron radiation
and CCD detector in Photon Factory in
[Back to top] Binding Potency of Peptide Fragments of Type
1 Collagen Cross-Linked N-Telopeptide Measured by an Enzyme-Linked Immunosorbant
Assay
Feng-Di
T. Lung , Chiu-Heng Chen1, Heuy-Yi Chen , Chien-Chung Liou , and Yen-Meng Liou
Osteoporosis
represents a major healthcare problem affecting elderly person. Urinary level
of the crosslinked N-telopeptide of type I collagen is a sensitive marker of
bone resorption. Ten overlapping peptides covering the N-telopeptide of alpha-2
type I collagen were synthesized, purified, and assayed for their relative
binding response to anti-type I collagen cross-linked N-telopeptide (NTX)
antibody by using a competitive-inhibition enzyme-linked immunosorbent assay
(ELISA). Peptides 1, 2, and 3, containing the N-terminal sequence of
Ntelopeptide, showed higher binding potency than peptides 4-10, suggesting that
these peptides may contain binding sites for anti-NTX antibodies, and can serve
as the lead for further preparation of their antibodies in order to develop
novel bioassays for monitoring the bone loss in humans.
[Back to top] Crystallization and preliminary X-ray
diffraction analysis of FKBP12 complexed with a novel neurotrophic ligand
Pengyun
Li , Liwei Wang , Yi Ding , Beili Wu ,
Cuiling Shu , Aihua Nie , Song Li , Beifen Shen
& Zihe Rao
A novel
neurotrophic ligand, (3R)-4-(p-Toluenesulfonyl)-1,4-thiazane-3-carboxylic
acid-L-Leucine ethyl ester, has been complexed with FKBP12 and crystallized
using the hanging-drop vapor-diffusion method. Crystals belong to P21 space
group, with unit cell parameters a=41.2, b=29.6, c=41.5Å, b=114.0°. The
crystals diffract to 1.8 Å resolution limit.