Biomedical subjects

Martin Malmsten

Publications and source records attributed to Martin Malmsten.

18 recordsLinked to original sources

PubMed2006

Effects of topology, length, and charge on the activity of a kininogen-derived peptide on lipid membranes and bacteria.

Effects of topology, length, and charge on peptide interactions with lipid bilayers was investigated for variants of the human kininogen-derived peptide HKH20 (HKHGHGHGKHKNKGKKNGKH) by ellipsometry, CD, fluorescence spectroscopy, and z-potential measurements. The peptides display primarily random coil conformation in buffer and at lipid bilayers, and their lipid interaction is dominated by electrostatics, the latter evidenced by higher peptide adsorption and resulting membrane rupture for an anionic than for a zwitterionic membrane, as well as by strongly reduced adsorption and membrane rupture at high ionic strength. At sufficiently high peptide charge density, however, electrostatic interactions contribute to reducing the peptide adsorption and membrane defect formation. Truncating HKH20 into overlapping 10 amino acid peptides resulted in essentially eliminated membrane rupture and in a reduced amount peptide charges pinned at the lipid bilayer. Finally, cyclic HKH20 was found to be less efficient than the linear peptide in causing liposome rupture, partly due to a lower adsorption. Analogous results were found regarding bactericidal effects.

Adsorption↗

PubMed2006

Preservation of antimicrobial properties of complement peptide C3a, from invertebrates to humans.

The human anaphylatoxin peptide C3a, generated during complement activation, exerts antimicrobial effects. Phylogenetic analysis, sequence analyses, and structural modeling studies paired with antimicrobial assays of peptides from known C3a sequences showed that, in particular in vertebrate C3a, crucial structural determinants governing antimicrobial activity have been conserved during the evolution of C3a. Thus, regions of the ancient C3a from Carcinoscorpius rotundicauda as well as corresponding parts of human C3a exhibited helical structures upon binding to bacterial lipopolysaccharide permeabilized liposomes and were antimicrobial against gram-negative and gram-positive bacteria. Human C3a and C4a (but not C5a) were antimicrobial, in concert with the separate evolutionary development of the chemotactic C5a. Thus, the results demonstrate that, notwithstanding a significant sequence variation, functional and structural constraints imposed on C3a during evolution have preserved critical properties governing antimicrobial activity.

Amino Acid Sequence↗

PubMed2006

The contact system--a novel branch of innate immunity generating antibacterial peptides.

Activation of the contact system has two classical consequences: initiation of the intrinsic pathway of coagulation, and cleavage of high molecular weight kininogen (HK) leading to the release of bradykinin, a potent proinflammatory peptide. In human plasma, activation of the contact system at the surface of significant bacterial pathogens was found to result in further HK processing and bacterial killing. A fragment comprising the D3 domain of HK is generated, and within this fragment a sequence of 26 amino acids is mainly responsible for the antibacterial activity. A synthetic peptide covering this sequence kills several bacterial species, also at physiological salt concentration, as effectively as the classical human antibacterial peptide LL-37. Moreover, in an animal model of infection, inhibition of the contact system promotes bacterial dissemination and growth. These data identify a novel and important role for the contact system in the defence against invasive bacterial infection.

Amino Acid Sequence↗

PubMed2006

Antifungal activity of C3a and C3a-derived peptides against Candida.

Antimicrobial peptides are generated during activation of the complement system [Nordahl et al. Proc. Natl. Acad. Sci. U. S. A. 2004, 101:16879-16884]. Here we show that the anaphylatoxin C3a exerts antimicrobial effects against the yeast Candida. Fluorescence microscopy and electron microscopy analysis demonstrated that C3a-derived peptides bound to the cell surface of Candida, and induced membrane perturbations and release of extracellular material. Various Candida isolates were found to induce complement degradation, leading to generation of C3a. Arginine residues were found to be critical for the antifungal and membrane breaking activity of a C3a-derived antimicrobial peptide, CNY21 (C3a; Cys57-Arg77). A CNY21 variant with increased positive net charge displayed enhanced antifungal activity. Thus, C3a-derived peptides can be utilized as templates in the development of peptide-based antifungal therapies.

Amino Acid Sequence↗

PubMed2006

Composition effect on peptide interaction with lipids and bacteria: variants of C3a peptide CNY21.

The effect of peptide hydrophobicity and charge on peptide interaction with model lipid bilayers was investigated for the C3a-derived peptide CNY21 by fluorescence spectroscopy, circular dichroism, ellipsometry, z-potential, and photon correlation spectroscopy measurements. For both zwitterionic and anionic liposomes, the membrane-disruptive potency for CNY21 variants increased with increasing net positive charge and mean hydrophobicity and was completely lost on elimination of all peptide positive charges. Analogous effects of elimination of the peptide positive net charge in particular were found regarding bacteria killing for both Pseudomonas aeruginosa and Bacillus subtilis. The peptides, characterized by moderate helix content both in buffer and when attached to the liposomes, displayed high adsorption for the net positively charged peptide variants, whereas adsorption was non-measurable for the uncharged peptide. That electrostatically driven adsorption represents the main driving force for membrane disruption in lipid systems was also demonstrated by a drastic reduction in both liposome leakage and peptide adsorption with increasing ionic strength, and this salt inactivation can be partly avoided by increasing the peptide hydrophobicity. This increased electrolyte resistance translates also to a higher antibacterial effect for the hydrophobically modified variant at high salt concentration. Overall, our findings demonstrate the importance of the peptide adsorption and resulting peptide interfacial density for membrane-disruptive effects of these peptides.

Adsorption↗

PubMed2006

Visualizing the interaction between poly-L-lysine and poly(acrylic acid) microgels using microscopy techniques: effect of electrostatics and peptide size.

The interaction between lightly cross-linked poly(acrylic acid) (pAA) microgels (50-150 microm in diameter) and poly-L-lysine (pLys) was studied as a function of pH, ionic strength, peptide size, and concentration. The swelling response and distribution of polypeptides within microgel particles was monitored by micromanipulator-assisted light microscopy and confocal laser scanning microscopy, while binding isotherms of pLys in the microgels were determined spectrophotometrically. Conformational changes of pLys were investigated by circular dichroism. The molecular weight of pLys was found to influence the degree of peptide-induced microgel deswelling, largely due to limitation of peptides larger than the effective network mesh size to penetrate the entire gel. Large peptides were concentrated within a surface layer of the gel particles, and at low ionic strength this dense surface layer was shown to act as a largely steric barrier for further penetration of compounds into the gel core. Small peptides, however, distributed evenly throughout the microgel particles and were able to create large microgel volume reductions. The deswelling of microgels increased with decreasing pH, while the uptake of pLys was significantly reduced at low pH. The effect of ionic strength on the interactions of pLys and oppositely charged pAA microgels was moderate and only pronounced for deswelling of gels at high pH. A significant increase in the alpha-helix content of pLys interacting with the oppositely charged microgels was observed for high molecular weight peptides, and the extent of alpha-helix formation was as expected more pronounced at high pH, i.e., at high charge density of the microgels but reduced charge density of the peptides.

Acrylic Resins↗

PubMed2006

Effect of peptide length on the interaction between consensus peptides and DOPC/DOPA bilayers.

The effect of peptide length and electrostatics on the interaction between Cardin motif peptides and lipid membranes was investigated for (AKKARA)(n) (n = 1-4) and (ARKAAKKA)(n) (n = 1-3) peptides (A, K, and R refer to alanine, lysine, and arginine, respectively) by fluorescence spectroscopy, circular dichroism, ellipsometry, z potential, and photon correlation spectroscopy measurements. The effect of the peptides regarding leakage induction of both zwitterionic and anionic liposomes increased with increasing peptide length, as did the peptide-induced killing of Enterococcus faecalis and Bacillus subtilis bacteria. The peptides, characterized by a random coil conformation both in buffer and when attached to the liposomes (helix content less than 20%), displayed an increased adsorption with increasing peptide length, and plateau adsorption for the longest peptides corresponded to 1 peptide per 65 and 17 lipid molecules for zwitterionic and anionic membranes, respectively. Control experiments with uncharged peptide analogues as well as experiments at high excess electrolyte concentration showed that peptide charges are important both for peptide adsorption and leakage induction. These observations, together with observations of the liposome z potential at different peptide additions as well as a comparison between the results for zwitterionic and anionic liposomes, suggest that electrostatically affected local packing effects are crucial for the action of these peptides, although pore formation such as that observed for many AMPs cannot be excluded at present.

Amino Acid Sequence↗

PubMed2006

Bacterial killing by heparin-binding peptides from PRELP and thrombospondin.

Antimicrobial peptides (AMP) are effector molecules of the innate immune system. A cross-functionality exists between AMPs and heparin-binding peptides. Here, we show that the peptides QPTRRPRPGTGPGRRPRPRPRP (QPT22), derived from proline arginine-rich end leucine-rich repeat protein (PRELP) and KRFKQDGGWSHWSPWSS (KRF17) from thrombospondin exert antimicrobial activities against the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, as well as against the Gram-positive Bacillus subtilis. Analysis by fluorescence microscopy demonstrated that QPT22 and KRF17 bind to bacterial membranes. Particularly QPT22 was found to induce membrane leakage of liposomes. The peptides displayed no hemolytic activities, nor did they exert membrane permeabilising effects on human epithelial cells. Additional peptides derived from heparin-binding regions of laminin, vitronectin, and fibronectin exerted similar antibacterial effects. Several peptides also showed activity against Staphylococcus aureus. Thus, the data disclose a novel antimicrobial activity of heparin-binding regions of matrix glycoproteins. The findings can be utilized in the development of novel AMPs for therapeutic use.

Amino Acid Sequence↗

PubMed2006

In silico identification and biological evaluation of antimicrobial peptides based on human cathelicidin LL-37.

Bacterial lipopolysaccharides (LPS) are important triggers of the widespread inflammatory response, which contributes to the development of multiple organ failure during sepsis. The helical 37-amino-acid-long human antimicrobial peptide LL-37 not only possesses a broad-spectrum antimicrobial activity but also binds and neutralizes LPS. However, the use of LL-37 in sepsis treatment is hampered by the fact that it is also cytotoxic. To find a less toxic analog of LL-37, we used in silico analysis to identify amphipathic helical regions of LL-37. A 21-amino-acid fragment (GKE) was synthesized, the biological actions of which were compared to those of two equally long peptides derived from the N and C termini of LL-37 as well as native LL-37. GKE displayed antimicrobial activity against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Candida albicans, and Candida parapsilosis that was similar to or even stronger than LL-37. GKE, as well as the equally long control peptides, attracted granulocytes in a fashion similar to that of LL-37, while only GKE was as potent as LL-37 in inhibiting LPS-induced vascular nitric oxide production. GKE caused less hemolysis and apoptosis in human cultured smooth muscle cells than LL-37. In summary, we have identified an active domain of LL-37, GKE, which displays antimicrobial activity in vitro and LPS-binding activity similar to those of LL-37 but is less toxic. GKE therefore holds promise as a template for the development of peptide antibiotics for the treatment of sepsis.

Amino Acid Sequence↗

PubMed2005

Model cellulose films exposed to H. insolens glucoside hydrolase family 45 endo-cellulase--the effect of the carbohydrate-binding module.

The effects of enzyme structure and activity on the degradation of model cellulose substrates were investigated by ellipsometry for the cellulase Humicola insolens GH45. The inactive variant D10N was found to adsorb at the cellulose surface but also to be incorporated into the cellulose films to an extent that depended on pH. For the native protein, the initial adsorption monitored for the inactive variant D10N was followed by enzyme-mediated degradation of the cellulose films. Again, a dependence on pH was found, such that higher pH resulted in slower enzymatic degradation. Removing the carbohydrate-binding module eliminated this pH dependence but also resulted in a decreased adsorption to the cellulose surface, and in a decreased net catalytic effect.

Ascomycota↗

PubMed2005

Enzymatic degradation of model cellulose films.

Enzymatic degradation of model cellulose films prepared by a spin-coating technique was investigated by ellipsometry. The cellulose films were prior to degradation characterized by ellipsometry, contact angle measurements, ESCA (electron spectroscopy for chemical analysis) and AFM (atomic force microscopy). At enzyme addition to preformed cellulose films an initial adsorption was observed, which was followed by a total interfacial mass decrease due to enzymatic degradation of the cellulose films. The degradation rate was found to be constant during an extended time of hours, whereafter the degradation leveled off. In parallel to the decreased interfacial mass, the cellulose degradation resulted in a thinner and more dilute interfacial film. At long degradation times, however, there was an expansion of the cellulose film. The enzyme concentration affected the degradation rate significantly, with a faster degradation at a higher enzyme concentration. The effects of pH, temperature, ionic strength and stirring rate in the cuvette were also investigated.

Adsorption↗

PubMed2004

Activation of the complement system generates antibacterial peptides.

The complement system represents an evolutionary old and significant part of the innate immune system involved in protection against invading microorganisms. Here, we show that the anaphylatoxin C3a and its inactivated derivative C3a-desArg are antibacterial, demonstrating a previously unknown direct antimicrobial effect of complement activation. The C3a peptide, as well as functional epitopes in the sequence, efficiently killed the Gram-negative bacteria Escherichia coli, Pseudomonas aeruginosa, and the Gram-positive Enterococcus faecalis. In mice, a C3a-derived peptide suppressed infection by Gram-positive Streptococcus pyogenes bacteria. Fluorescence and electron microscopy demonstrated that C3a binds to and induces breaks in bacterial membranes. C3a was also found to induce membrane leakage of liposomes. These findings provide an interesting link between the complement system and antimicrobial peptides, which are two important branches of innate immunity.

Amino Acid Sequence↗

PubMed2004

An ellipsometry-based Alzheimer plaque mimic: Effect of beta-amyloid, lipoprotein identity and apolipoprotein E isoform.

Ca2+-induced deposition of low-density lipoprotein (LDL), intermediate-density lipoprotein (IDL), and high-density lipoprotein (HDL) at proteoheparan sulfate-modified surfaces was investigated as a function of beta-amyloid (Abeta) presence and apolipoprotein E isoform. Presence of beta-amyloid resulted in an increased deposition, as did the E4/E4 isoform compared to the corresponding E3/E3 isoform. The results are compatible with findings reported in literature on plaque formation in Alzheimer's disease, and suggest that, although simplistic, the present model system may have some potential in biosensor studies of Alzheimer plaque formation.

Alzheimer Disease↗

PubMed2004

[Inhibition of arteriosclerotic plaque development by garlic].

OBJECTIVE: In an in vitro biosensor model (PCT/EP 97/05212), the interplay between different lipoproteins in arteriosclerotic nanoplaque formation, as well as aqueous garlic extract (0.2-5.0 g/l from LI 111 powder) as a possible candidate drug against arterio/atherosclerosis were tested within the frame of a high throughput screening. METHODS: The processes described below were studied by ellipsometric techniques quantifying the adsorbed amount (nanoplaque formation) and layer thickness (nanoplaque size). A thorough description of the experimental setup has been given previously. RESULTS: Proteoheparan sulfate (HS-PG) adsorption to hydrophobic silica was monoexponential and after approximately 30 min constant. The LDL plasma fraction (100 mg/dl) from a healthy probationer showed beginning arteriosclerotic nanoplaque formation already at a normal blood Ca2+ concentration, with a strong increase at higher Ca2+ concentrations. Aqueous garlic extract (GE), preferably in a concentration of 1 g/l, applied acutely in the experiment, markedly slowed down this process of ternary aggregational nanoplaque complexation at all Ca2+ concentrations used. In a normal blood Ca2+ concentration of 2.52 mmol/l, the garlic induced reduction of nanoplaque formation and molecular size amounted to 14.8 % and 3.9%, respectively, as compared to the controls. Furthermore, after ternary complex build-up, GE similar to HDL, was able to reduce nanoplaque formation and size. The incubation time for HDL and garlic was only 30 min each in these experiments. Nevertheless, after this short time the deposition of the ternary complex decreased by 6.2% resp. 16.5%, i.e. the complex aggregates were basically resolvable. CONCLUSIONS: These experiments clearly proved that garlic extract strongly inhibits Ca2+ binding to HS-PG. In consequence, the formation of the ternary HS-PG/LDL/Ca2+ complex, initially responsible for the 'nanoplaque' composition and ultimately for the arteriosclerotic plaque generation, is decisively blunted.

Adsorption↗

PubMed2003

Protein-polyelectrolyte cluster formation and redissolution: a Monte Carlo study.

Aqueous solutions of proteins and oppositely charged polyelectrolytes were studied at different polyelectrolyte chain length, ionic strength, and protein-protein interaction potential as a function of the polyelectrolyte concentration. One of the protein models used represented lysozyme in aqueous environment. The model systems were solved by Monte Carlo simulations, and their properties were analyzed in terms of radial distribution functions, structure factors, and cluster composition probabilities. In the system with the strongest electrostatic protein-polyelectrolyte interaction the largest clusters were formed near or at equivalent amount of net protein charge and polyelectrolyte charge, whereas in excess of polyelectrolyte a redissolution appeared. Shorter polyelectrolyte chains and increased ionic strength lead to weaker cluster formation. An inclusion of nonelectrostatic protein-protein attraction promoted the protein-polyelectrolyte cluster formation.

Cluster Analysis↗

PubMed2003

Adsorption from black tea and red wine onto in vitro salivary pellicles studied by ellipsometry.

The adsorption of black tea and red wine components onto a pellicle-like protein layer formed in vitro by adsorption from whole unstimulated saliva on hydroxyapatite discs were studied by in situ ellipsometry. It was found that components from black tea readily adsorbed to the pellicle. Subsequent exposure to saliva led to further adsorption of salivary components to give an overall increase in the amounts adsorbed. The amounts adsorbed increased still further following a third tea and saliva exposure. Components of red wine gave significantly greater amounts of adsorption to the pellicle than black tea. The adsorption of components of black tea gave a concomitant increase in colour or stain as measured by a reflectance chromameter. In all cases, the black tea- and red wine-modified pellicles were not eluted by either phosphate buffer or sodium dodecyl sulphate (SDS) rinses. Thus, black tea and red wine components have been shown to have a profound effect on in vitro pellicle maturation, causing thickened layers of stained material to build up, which are not readily removed.

Adsorption↗

PubMed2001

A Model Substrate for Ellipsometry Studies of Lipoprotein Deposition at the Endothelium.

Endothelial cell layers from umbilical cords were grown on methylated silica surfaces. Fluorescence microscopy showed this layer to be confluent and to consist of living cells. Initial ellipsometry measurements were performed to illustrate both the stability of the model surface in ellipsometry measurements and the Ca(2+)-dependent binding of lipoproteins at the cell-based substrate. The present substrate holds promise as a model substrate for in vitro investigations of lipoprotein deposition at the endothelium surface under close to in vivo conditions. Copyright 2001 Academic Press.

Journal Article↗

PubMed2001

Adsorption of Poly(ethylene oxide)-Poly(lactide) Copolymers. Effects of Composition and Degradation.

The effect of chemical degradation of two diblock copolymers of poly(ethylene oxide) (E) and poly(lactide) (L), E(39)L(5) and E(39)L(20), on their adsorption at silica and methylated silica was investigated with in situ ellipsometry. Steric stablization of polystyrene dispersions was investigated in relation to degradation. Hydrolysis of the poly(lactide) block of the copolymers was followed at different temperatures and pH by using HPLC to measure the occurrence of lactic acid in solution. The block copolymers were quite stable in pH-unadjusted solution at low temperature, whereas degradation was facilitated by increasing temperature or lowering of the pH. Lower degradation rates of E(39)L(20) where observed at low temperature in comparison with those of E(39)L(5), whereas the degradation rates of the copolymers were quantitatively similar at high temperature. The adsorption of the copolymers at methylated silica substrates decreased with increasing degree of degradation due to the reduction in the ability of hydrophobic block to anchor the copolymer layer at the surface. At silica the adsorption initially increased with increasing degradation, particularly for E(39)L(20) due to deposition of aggregates onto the surface. After extensive degradation the adsorption of the copolymers at both silica and methylated silica resembled that of the corresponding poly(ethylene oxide) homopolymer. Overall, it was found that the eventual reduction in adsorption occurred at a lower degree of degradation for E(39)L(5) than for E(39)L(20). Mean-field calculations showed a reduced anchoring for the block copolymers with decreasing poly(lactide) block length at hydrophobic surfaces. In accordance with this finding, it was observed that polystyrene dispersions were stabilized by E(39)L(20) or E(39)L(5) in a way that depended on both the lactide block length and the degree of degradation. Upon degradation of the hydrophobic block, stabilization of the polystyrene dispersions was maintained initially, but eventually degradation resulted in destabilization. The average residual copolymer concentration required for stabilization of the polystyrene dispersions was much higher than the corresponding concentration of intact copolymer required for stabilization. Copyright 2001 Academic Press.

Journal Article↗