共查询到20条相似文献,搜索用时 15 毫秒
1.
Jelokhani-Niaraki M Hodges RS Meissner JE Hassenstein UE Wheaton L 《Biophysical journal》2008,95(7):3306-3321
To investigate the mechanism of interaction of gramicidin S-like antimicrobial peptides with biological membranes, a series of five decameric cyclic cationic β-sheet-β-turn peptides with all possible combinations of aromatic D-amino acids, Cyclo(Val-Lys-Leu-D-Ar1-Pro-Val-Lys-Leu-D-Ar2-Pro) (Ar ≡ Phe, Tyr, Trp), were synthesized. Conformations of these cyclic peptides were comparable in aqueous solutions and lipid vesicles. Isothermal titration calorimetry measurements revealed entropy-driven binding of cyclic peptides to POPC and POPE/POPG lipid vesicles. Binding of peptides to both vesicle systems was endothermic—exceptions were peptides containing the Trp-Trp and Tyr-Trp pairs with exothermic binding to POPC vesicles. Application of one- and two-site binding (partitioning) models to binding isotherms of exothermic and endothermic binding processes, respectively, resulted in determination of peptide-lipid membrane binding constants (Kb). The Kb1 and Kb2 values for endothermic two-step binding processes corresponded to high and low binding affinities (Kb1 ≥ 100 Kb2). Conformational change of cyclic peptides in transferring from buffer to lipid bilayer surfaces was estimated using fluorescence resonance energy transfer between the Tyr-Trp pair in one of the peptide constructs. The cyclic peptide conformation expands upon adsorption on lipid bilayer surface and interacts more deeply with the outer monolayer causing bilayer deformation, which may lead to formation of nonspecific transient peptide-lipid porelike zones causing membrane lysis. 相似文献
2.
Bio-deposition of a calcium carbonate layer on degraded limestone by Bacillus species 总被引:4,自引:0,他引:4
Dick J De Windt W De Graef B Saveyn H Van der Meeren P De Belie N Verstraete W 《Biodegradation》2006,17(4):357-367
To obtain a restoring and protective calcite layer on degraded limestone, five different strains of the Bacillus sphaericus group and one strain of Bacillus lentus were tested for their ureolytic driven calcium carbonate precipitation. Although all the Bacillus strains were capable of depositing calcium carbonate, differences occurred in the amount of precipitated calcium carbonate on agar plate colonies. Seven parameters involved in the process were examined: calcite deposition on limestone cubes, pH increase, urea degrading capacity, extracellular polymeric substances (EPS)-production, biofilm formation, ζ-potential and deposition of dense crystal layers. The strain selection for optimal deposition of a dense CaCO3 layer on limestone, was based on decrease in water absorption rate by treated limestone. Not all of the bacterial strains were effective in the restoration of deteriorated Euville limestone. The best calcite precipitating strains were characterised by high ureolytic efficiency, homogeneous calcite deposition on limestone cubes and a very negative ζ-potential. 相似文献
3.
G. E. Dobretsov T. I. Syreishchikova Yu. A. Gryzunov N. V. Smolina A. A. Komar 《Biophysics》2010,55(2):182-187
A study was made of the binding of a fluorescent probe K-35 (N-(carboxyphenyl)imide of 4-(dimethylamino)naphthalic acid),
used as an indicator of albumin structural changes in pathology, to human serum albumin (HSA). Based on the data on the fluorescence
decay of the probe, four types of site of K-35 binding to HSA have been recognized, which differ in fluorescence decay time
(τ) and binding constant (K). Probe molecules bound to the first type of site have a decay time of 8–10 ns; this value corresponds to a high fluorescence
quantum yield of about 0.7. These sites have a maximal binding constant, K
1 = 5 · 104 M−1. The τ2 of the second type of site is close to 3.6 ns and K
2 = 1 · 104 M−1, which is much lower than K
1; however, the number of these sites is several times greater. The number of sites of the third type and the binding constant
are close to those of the second type, but the decay time τ3 is 1 ns, which is significantly lower than τ2. The binding of K-35 to sites of the second and the third types is characterized by a positive cooperativity. Their properties
are similar but not completely identical. The total number of sites of these three types is about two per one HSA molecule.
There are also one-two sites of the fourth type where bound K-35 molecules have a very short decay time τ4 ≪ 1, i.e., are virtually nonfluorescent, and K
4 = 1 · 104 M−1. The major contribution to the steady-state fluorescence is made by probe molecules bound to sites of the first and second
types. As a rule, the concentration of albumin binding sites in blood is significantly higher than the concentration of metabolites
and xenobiotics transferred by albumin. Therefore, the metabolite—or the probe in these experiments—is distributed among different
sites in accordance with their K
i
n
i
values (n
i
is the number of sites of the i-th type per albumin molecule). The low occupancy of the sites results in an approximately equal number of K-35 molecules
bound to different sites of types 1, 2, and 3. The competition of K-35 with phenylbutazone, a marker of the albumin drug-binding
site I, allows one to suggest that the K-35 site of the first type is localized exactly in the drug site I region, while the
sites of the second and third types are close to it. 相似文献
4.
In this work, fluorescence spectroscopy in combination with circular dichroism spectroscopy and molecular modeling was employed
to investigate the binding of 10-hydroxycamptothecin (HCPT) to human serum albumin (HSA) under simulative physiological conditions.
The experiment results showed that the fluorescence quenching of HSA by HCPT was a result of the formation of HCPT–HSA complex.
The corresponding association constants (K
a) between HCPT and HSA at four different temperatures were determined according to the modified Stern–Volmer equation. The
results of thermodynamic parameters ΔG, ΔH, and ΔS indicated that hydrogen bonds and van der Waals forces played major roles for HCPT–HSA association. Site marker competitive
displacement experiment indicated that the binding of HCPT to HSA primarily took place in sub-domain IIA (site I). Molecular
docking study further confirmed the binding mode and the binding site obtained by fluorescence and site marker competitive
experiments. The conformational investigation showed that the presence of HCPT decreased the α-helical content of HSA and
induced the slight unfolding of the polypeptides of protein, which confirmed some micro-environmental and conformational changes
of HSA molecules. 相似文献
5.
Cibacron Blue F3G-A, a probe used to monitor nucleotide binding domains in enzymes, inhibited sheep liver 5,10-methylenetetrahydrofolate
reductase competitively with respect to 5-methyltetrahydrofolate and NADPH. TheK
i values obtained by kinetic methods and theK
d value for the binding of the dye to the enzyme estimated by protein fluorescence quenching were in the range 0.9–1.2 μM.
Another triazine dye, Procion Red HE-3B interacted with the enzyme in an essentially similar manner to that observed with
Cibacron Blue F3G-A. These results as well as the interaction of the dye with the enzyme monitored by difference spectroscopy
and intrinsic protein fluorescence quenching methods indicated that the dye was probably interacting at the active site of
the enzyme by binding at a hydrophobic region. 相似文献
6.
Tryptophan (Trp) surface topography of the red- and far-red-absorbing forms of phytochrome (Pr, Pfr) ofAvena sativa L. has been investigated by analyzing quenching of the two components of Trp fluorescence decay, in order to understand the
differences in the two forms at the molecular level. Stern-Volmer kinetic analysis of the quenching data for two cationic
surface quenchers, Cs+ and Tl+, showed strong quenching of the short component of the Pr fluorescence (Stern-Volmer constants,K
sv
, 27.2 and 21.4 M−1, respectively) relative to that of Pfr fluorescenceK
sv
, 10.4 and 12.3 M−1, respectively). The long component of the Trp fluorescence was quenched differentially by Cs+ and Tl+, withK
sv
of 9.0 and 19.8 M−1, respectively, for the Pr fluorescence andK
sv
of 13.7 and 8.7 M−1, respectively, for the Pfr fluorescence. The results indicate that the phytochrome Trp residues with short fluorescence lifetime
are more accessible to the cationic surface quenchers than those with long fluorescence lifetime. The data, taken together
with our earlier study (Singh et al. 1988, Biochim, Biophys. Acta936, 395–405), indicate that most, if not all the ten Trp residues of phytochrome, are fluorescent and exist in distinct groups
differing in their topography and microenvironment, and the peptide segment containing Trp-774 and Trp-778 within the 55-kilodalton
C-terminal domain of phytochrome also undergoes a subtle alteration in its surface topography during Pr→Pfr phototransformation.
This paper is dedicated to Professor Hans Mohr in commemoration of his 60th birthday 相似文献
7.
We suggest a novel approach for direct optical microscopy observation of DNA interaction with the bilayers of giant cationic
liposomes. Giant unilamellar vesicles, about 100 μm in diameter, made of phosphatidylcholines and up to 33 mol% of the natural
bioactive cationic amphiphile sphingosine, were obtained by electroformation. “Short” DNAs (oligonucleotide 21b and calf thymus
250 bp) were locally injected by micropipette to a part of the giant unilamellar vesicle (GUV) membrane. DNAs were injected
native, as well as marked with a fluorescent dye. The resulting membrane topology transformations were monitored in phase
contrast, while DNA distribution was followed in fluorescence. We observed DNA-induced endocytosis due to the DNA/lipid membrane
local interactions and complex formation. A characteristic minimum concentration (C
endo) of d-erythro-sphingosine (Sph+) in the GUV membrane was necessary for the endocytic phenomenon to occur. Below C
endo, only lateral adhesions between neighboring vesicles were observed upon DNA local addition. C
endo depends on the type of zwitterionic (phosphocholine) lipid used, being about 10 mol% for DPhPC/Sph+ GUVs and about 20 mol% for SOPC/Sph+ or eggPC/Sph+ GUVs. The characteristic sizes and shapes of the resulting endosomes depend on the kind of DNA, and initial GUV membrane
tension. When the fluorescent DNA marker dye was injected after the DNA/lipid local interaction and complex formation, no
fluorescence was detected. This observation could be explained if one assumes that the DNA is protected by lipids in the DNA/lipid
complex, thereby inaccessible for the dye molecules. We suggest a possible mechanism for DNA/lipid membrane interaction involving
DNA encapsulation within an inverted micelle included in the lipid membrane. Our model observations could help in understanding
events associated with the interaction of DNA with biological membranes, as well as cationic liposomes/DNA complex formation
in gene transfer processes.
Received: 18 April 1998 / Revised version: 6 August 1998 / Accepted: 7 August 1998 相似文献
8.
The aim of this work was to study the effect of antimicrobial peptides: divergicin M35 and nisin A on Listeria monocytogenes LSD 530 potassium (K+) channels: ATP-sensitive (KATP), calcium-activated (BKCa), and depolarization-activated (Kv) types. Increase on K+ efflux and inhibition of cellular growth were observed after adding K+ channel activators pinacidil, NS1619, and cromakalim to divergicin M35. Increase in K+ efflux from log-phase cells was about 18 ± 1.1, 11 ± 0.63, and nmol mg−1 of cell dry weight (CDW) for pinacidil and NS1619, respectively, over the efflux obtained with divergicin M35 alone. Increases
in K+ efflux obtained by adding the same K+ channel activators to nisin A fit a completely different profile. Divergicin M35 activates K+ channels, particularly of the Kv and BKCa types and to a lesser extent the KATP type, causing K+ efflux and consequently cell death. 相似文献
9.
In the muscle wall of the earthworm Lumbricus terrestris, with the aid of fluorescent endocytotic dyes FM1-43, FM2-10, and FM4-64, there are revealed fluorescent spots 1–2 μm in
diameter that represent clusters of “synaptic boutons.” Application takes place onto ganglia of the abdominal nerve chain
of the Dil membrane probe capable of translocation by axoplasmic transport; the subsequent (next day) staining of nerve structures
with the endocytotic marker FM4-64 showed the complete superposition of fluorescence of these dyes fluorescing in different
specter areas. The fluorescent marker DiBAC4(3) revealed an enhancement of fluorescence of nerve elements with increase of K+ concentration in the extracellular medium. Use of FM2-10 showed that, the higher the K+ content in solution and, accordingly, the nerve cell depolarization, the faster the release of the marker and, on the contrary,
the slower the process in the absence of K+ in the medium. In the Ca2+-free solution and in the presence of the Ca2+ chelator BAPTA or BAPTA-AM, there uptake and release of FM2-10 are blocked, but only after preliminary 40-min incubation
in such solution. In clusters of synaptic boutons, exo- and endocytosis processes take place that are also preserved under
conditions of rest. This vesicular cycle depends on the membrane potential of nerve structures and on the content of K+ and Ca2+ in the medium, the calcium sensor working most likely by the “all or nothing” principle. 相似文献
10.
Surfactin is a lipopeptide produced by Bacillus subtilis which possesses antimicrobial activity. We have studied the leakage and lysis of POPC vesicles induced by surfactin using
calcein fluorescence de-quenching, isothermal titration calorimetry and 31P solid state NMR. Membrane leakage starts at a surfactin-to-lipid ratio in the membrane, R
b ≈ 0.05, and an aqueous surfactin concentration of C
Sw ≈ 2 μM. The transient, graded nature of leakage and the apparent coupling with surfactin translocation to the inner leaflet
of the vesicles, suggests that this low-concentration effect is due to a bilayer-couple mechanism. Different permeabilization
behaviour is found at R
b ≈ 0.15 and attributed to surfactin-rich clusters, which can induce leaks and stabilize them by covering their hydrophobic
edges. Membrane lysis or solubilization to micellar structures starts at R
bsat = 0.22 and C
Sw = 9 μM and is completed at R
msol = 0.43 and C
Sw = 11 μM. The membrane–water partition coefficient of surfactin is obtained as K = 2 × 104 M−1. These data resolve inconsistencies in the literature and shed light on the variety of effects often referred to as detergent-like
effects of antibiotic peptides on membranes. The results are compared with published parameters characterizing the hemolytic
and antibacterial activity.
Dedicated to Prof. K. Arnold on the occasion of his 65th birthday. 相似文献
11.
Effects of Cations and PH on Antimicrobial Activity of Thanatin and s-Thanatin Against Escherichia coli ATCC25922 and B. subtilis ATCC 21332 总被引:1,自引:0,他引:1
This study analyzes the in vitro effects of cations and pH on antimicrobial activity of thanatin and s-thanatin against Escherichia coli ATCC25922 and B. subtilis ATCC21332. Thanatin and s-thanatin were synthesized by the solid-phase method using a model 432A synthesizer. The bacterial
strains tested included two antibiotic-susceptible strains of Escherichia coli ATCC25922 and B. subtilis ATCC21332. Susceptibility determinations were carried out either in a variety of cation concentrations or in pH conditions
from pH 5 to pH 8. NaCl or KCl was added to the media to final concentrations of 0, 10, 50, 100, 200, and 500 mM, whereas
CaCl2 and MgCl2 were added to the media to final concentrations of 0, 1, 2, 5, 10, and 20 mM. The antimicrobial activity of thanatin and
s-thanatin against Escherichia coli ATCC25922 and B. subtilis ATCC21332 decreased, as indicated by the increasing minimal inhibitory concentrations (MICs) of both peptides with increasing
concentrations of Na+/K+/Ca2+/Mg2+. Both peptides lost their activities at 500 mM Na+/K+ but retained them at 20 mM Ca2+/Mg2+. Both peptides have MICs that are not significantly different at a variety of pH levels, with the antimicrobial activity
slightly higher in neutral or slightly basic media than under acidic conditions. The antimicrobial peptides thanatin and s-thanatin,
which have an anti-parallel β-sheet constrained by disulfide bonds, were salt sensitive against both Gram-positive and Gram-negative
pathogens in vitro. Determining the reason why the thanatins are salt sensitive would be useful to provide an understanding
of how thanatin and s-thanatin kill bacteria. Futher investigation of the antimicrobial properties of these peptides is warranted.
G. Wu and J. Ding contributed equally to this article. 相似文献
12.
The protein dipole moment is a low-resolution parameter that characterizes the second-order charge organization of a biomolecule.
Theoretical approaches to calculate protein dipole moments rely on pK
a values, which are either computed individually for each ionizable residue or obtained from model compounds. The influence
of pK
a shifts are evaluated first by comparing calculated and measured dipole moments of β-lactoglobulin. Second, calculations are made on a dataset of 66 proteins from the Protein Data Bank, and average differences
are determined between dipole moments calculated with model pK
as, pK
as derived using a Poisson–Boltzmann approach, and empirically-calculated pK
as. Dipole moment predictions that neglect pK
a shifts are consistently larger than predictions in which they are included. The importance of pK
a shifts are observed to vary with protein size, internal permittivity, and solution pH. 相似文献
13.
The present study is aimed at developing and exploring the use of pectin in suppression of agglomeration of ciprofloxacinloaded
human serum albumin (HSA) nanoparticles. The HSA-pectin nanoparticles loaded with ciprofloxacin were prepared by the pH-coacervation
method, and various physicochemical parameters such as particle size, morphology, ζ-potential, electrolyte-induced flocculation,
pH-dependent ζ-potential, drug loading, in vitro drug release, and stability of nanoparticles, were evaluated. The size of
the HSA-pectin nanoparticles (F3) was found to be 180 to 290 nm. The HSA nanoparticles were modified with pectin when the
critical flocculation concentration of nanoparticles in Na2SO4 solution was increased from 0.3 M to 0.9 M. The isoelectric points of the formed nanoparticles were found to be relatively
lower between pH values 3 and 6. Pectin may be used as a pharmaceutical additive for the suppression of particle agglomeration
in HSA nanoparticles, and the effect may be attributed to the pectin segments present on the surface of nanoparticles.
Published: March 2, 2007 相似文献
14.
On binding toVicia faba lectin, the fluorescence of 4-methylumbelliferyl-α-D-glucoPyranoside was quantitatively quenched showing that the interaction
of 4-methylumbelliferyl-α-D-glucoPyranoside took Place in a binding environment. The binding of the fluorescent sugar was
saccharide sPecific as evidenced by the reversal of 4-methylumbelliferyl-α-D-glucoPyranoside fluorescence quenching by D-fructose.
The association constant,K
a, values for the 4-methylumbelliferyl-α-D-glucoPyranoside was determined by comPetition study emPloying reversal of fluorescence
quenching of 4-methylumbelliferyl-α-D-glucoPyranoside by D-fructose. TheK
a value obtained for D-fructose was 1.07 ±0.03 X 104 M-1 and for 4-methylumbelliferyl-α-D-glucoPyranoside was 1.60 ±0.05 X 104 M-1 at 15°C. TheK
a values of 2.51 ±0.06 X 104M-1, l.26 ±0.02 X 104 M-1 and 0.56 ±0.01 X 104M-1, resPectively at 10°, 20° and 30°C were obtained from the ChiPman equation. The relative fluorescence quenching, ΔF
a, at infinite concentration of the free saccharide sites ofVicia faba lectin [P′] was 93.5% at 30°C and the binding constant for 4-methylumbelliferyl-α-D-glucoPyranoside lectin interaction as derived by
Yank and Hanaguchi equation was 0.63 ±0.01 X 104M-1. 相似文献
15.
We have investigated the interaction of two peptides (ShB — net charge +3 and ShB:E12KD13K — net charge +7) derived from the NH2-terminal domain of the Shaker K+ channel with purified, ryanodine-modified, cardiac Ca2+-release channels (RyR). Both peptides produced well resolved blocking events from the cytosolic face of the channel. At a
holding potential of +60 mV the relationship between the probability of block and peptide concentration was described by a
single-site binding scheme with 50% saturation occurring at 5.92 ± 1.06 μm for ShB and 0.59 ± 0.14 nm for ShB:E12KD13K. The association rates of both peptides varied with concentration (4.0 ± 0.4 sec−1μm
−1 for ShB and 2000 ± 200 sec−1μm
−1 for ShB:E12KD13K); dissociation rates were independent of concentration. The interaction of both peptides was influenced by applied
potential with the bulk of the voltage-dependence residing in Koff. The effectiveness of the inactivation peptides as blockers of RyR is enhanced by an increase in net positive charge. As
is the case with inactivation and block of K+ channels, this is mediated by a large increase in Kon. These observations are consistent with the proposal that the conduction pathway of RyR contains negatively charged sites
which will contribute to the ion handling properties of this channel.
Received: 15 December 1997/Revised: 13 March 1998 相似文献
16.
S. A. M. Otman A. B. Pshenichnikova V. I. Shvets 《Applied Biochemistry and Microbiology》2012,48(2):200-205
Accelerating growth and increasing exopolysaccharide production in obligate methylotrophic bacterium Methylophilus quaylei were observed in the presence of C12–C18 fatty acids added to the growth media. Sodium oleate was the best growth factor. Based on data on the composition of the
free fatty acids fraction in the cells and the values of the ξ-potential and fluorescence anisotropy of whole cells, we suggested
that fatty acids were incorporated in the outer membrane of M. quaylei. 相似文献
17.
Summary Pararosaniline-Feulgen staining of cells in suspension produces nucleus- and chromatin-specific fluorescence as well as color.
Experiments were designed to test postulated reaction mechanisms responsible for the fluorescent staining with the nonfluorescent
pararosaniline. The reduction in fluorescent-staining intensity by pretreatment of cells with 2.2×10−2M K2S2O5 tends to rule out the alkysulfonic acid pathway; conditions favoring the formation of this intermediate reduce staining intensity.
The fluorescence enhancement, observed when cells stained in pararosaniline without K2S2O5 are post-treated with K2S2O5, suggests that there is an initial Schiff-base linkage between pararosaniline and an aldehyde of hydrolyzed DNA, and that
this linkage is stabilized in the presence of K2S2O5. Microspectrofluorometer measurements of cells stained at various pararosaniline concentrations in 2.2×10−2M K2S2O5, show that the fluorescence emission maximum ranges from about 627 nm at 3.1×10−3M pararosaniline to about 604 nm at 3.1×10−5M. All of the employed staining protocols appear to produce the same fluorescent product, perhaps a heterocyclic pyronin analog
formed from pararosaniline. Flow microfluorometric analysis of cells stained in suspension verified that the relative fluorescence
intensity represents relative DNA content. Staining at reduced pararosaniline concentration (3.1×10−4M) reduces the coefficient of variation of the flow microfluorometric histograms, showing that maximum quantitation does not
necessarily correlate with maximum staining intensity. 相似文献
18.
Halocidin was isolated from hemocytes, Halocynthia aurantium as a heterodimeric peptide consisting of two α-helical subunits, Hal15 and Hal18. Hal18 was shown to have antibacterial properties
against Bacillus subtilis (MLC = 15 μM) and Escherichia coli (MLC = 100 μM). The peptide was shown to produce stable monolayers, which were characteristic of α-helical peptides predicted
to orientate parallel to the surface of the interface. Constant area assays showed that Hal18 was surface active (4 μM) inducing
surface pressure changes >30 mN m−1 characteristic of membrane interactive peptides. The peptide induced stable surface pressure changes in monolayers that were
mimetic of B. subtilis membranes (circa 7 mN m−1) and E. coli membrane-mimics (circa 4 mN m−1). Hal18 inserted readily into zwitterionic DOPE and anionic DOPG monolayers inducing surface pressure changes circa 8 mN m−1 in both cases, providing evidence that interaction is not headgroup specific. Thermodynamic analysis of compression isotherms
showed that the presence of Hal18 destabilised B. subtilis membranes (ΔG
Mix > 0), which is in contrast to its stabilising effect on E. coli lipid extract implying the differential antimicrobial efficacy may be driven by lipid packing. 相似文献
19.
J. M. Bateman 《Bulletin of mathematical biology》1985,47(5):651-668
This work continues with an examination of capillary exchange models as operators, namely the operatorsO
k
andK
αk
relating extravascular and intravascular concentration to input for the Krogh cylinder model of a single capillary, a model
basic to many organ models. Fundamental algebraic and analytic properties are presented: the operators belong to a commutative
Banach algebra; an addition theorem holdsK
αk
+K
βk
=K
α+β,k
; the operatorK
αk
has an inverse;K
αk
-1
, (as an operator on LebesgueL
p
space or on the locally integrable functions); partial derivatives are given forK
αk
[f](t) andO
k
[f](t) (sensitivity functions); and inequalities are established for the derivatives. Dominance relations between model curves are
inferred. Error bound formulas are presented forK andO as bounds on ‖K
αk
f-K
βl
f‖
p
and ‖O
k
f-O
l
f‖
p
for allL
p
. Consequent limitations on relative errors are shown. The implications for operators on a finite time interval are deduced.
This work supported in part by PHS Grant Nos HL-19153 (SCOR and Pulmonary Vascular Disease) and HL-19370 at Vanderbilt University
Medical School. 相似文献
20.
The antibacterial peptide CM4 (ABP-CM4), isolated from Chinese Bombys mori, is a 35-residue cationic, amphipathic α-helical peptide that exhibits a broad range of antimicrobial activity. To explore
a new approach for the expression of ABP-CM4 in E. coli, the gene ABP-CM4, obtained by recursive PCR (rPCR), was cloned into the vector pET32a to construct a fusion expression plasmid.
The fusion protein Trx-CM4 was expressed in soluble form, purified by Ni2+-chelating chromatography, and cleaved by formic acid to release recombinant CM4. Purification of rCM4 was achieved by affinity
chromatography and reverse-phase HPLC. The purified of recombinant peptide showed antimicrobial activities against E. coli K12D31, Penicillium chrysogenum, Aspergillus niger and Gibberella saubinetii. According to the antimicrobial peptide database (http://aps.unmc.edu/AP/main.html), 116 peptides contain a Met residue, but
only 5 peptides contain the AspPro site, indicating a broader application of formic acid than CNBr in cleaving fusion protein.
The successful application to the expression of the ABP-CM4 indicates that the system is a low-cost, efficient way of producting
milligram quantities of ABP-CM4 that is biologically active. 相似文献