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Synthetic Evaluation of Standard and Microwave-Assisted Solid Phase Peptide Synthesis of a Long Chimeric Peptide Derived from Four Plasmodium falciparum Proteins.

ABSTRACT: An 82-residue-long chimeric peptide was synthesised by solid phase peptide synthesis (SPPS), following the Fmoc protocol. Microwave (MW) radiation-assisted synthesis was compared to standard synthesis using low loading (0.20 mmol/g) of polyethylene glycol (PEG) resin. Similar synthetic difficulties were found when the chimeric peptide was obtained via these two reaction conditions, indicating that such difficulties were inherent to the sequence and could not be resolved using MW; by contrast, the number of coupling cycles and total reaction time became reduced whilst crude yield and percentage recovery after purification were higher for MW radiation-assisted synthesis.

SUBMITTER: Varela YF

PROVIDER: S-EPMC6278645 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Synthetic Evaluation of Standard and Microwave-Assisted Solid Phase Peptide Synthesis of a Long Chimeric Peptide Derived from Four <i>Plasmodium falciparum</i> Proteins.

Varela Yahson F YF Vanegas Murcia Magnolia M Patarroyo Manuel Elkin ME

Molecules (Basel, Switzerland) 20181105 11

An 82-residue-long chimeric peptide was synthesised by solid phase peptide synthesis (SPPS), following the Fmoc protocol. Microwave (MW) radiation-assisted synthesis was compared to standard synthesis using low loading (0.20 mmol/g) of polyethylene glycol (PEG) resin. Similar synthetic difficulties were found when the chimeric peptide was obtained via these two reaction conditions, indicating that such difficulties were inherent to the sequence and could not be resolved using MW; by contrast, th ...[more]

PMID: 30400576

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Project description:BackgroundFully efficient vaccines against malaria pre-erythrocytic stage are still lacking. The objective of this dose/adjuvant-finding study was to evaluate the safety, reactogenicity and immunogenicity of a vaccine candidate based on a peptide spanning the C-terminal region of Plasmodium falciparum circumsporozoite protein (PfCS102) in malaria naive adults.Methodology and principal findingsThirty-six healthy malaria-naive adults were randomly distributed into three dose blocks (10, 30 and 100 microg) and vaccinated with PfCS102 in combination with either Montanide ISA 720 or GSK proprietary Adjuvant System AS02A at days 0, 60, and 180. Primary end-point (safety and reactogenicity) was based on the frequency of adverse events (AE) and of abnormal biological safety tests; secondary-end point (immunogenicity) on P. falciparum specific cell-mediated immunity and antibody response before and after immunization. The two adjuvant formulations were well tolerated and their safety profile was good. Most AEs were local and, when systemic, involved mainly fatigue and headache. Half the volunteers in AS02A groups experienced severe AEs (mainly erythema). After the third injection, 34 of 35 volunteers developed anti-PfCS102 and anti-sporozoite antibodies, and 28 of 35 demonstrated T-cell proliferative responses and IFN-gamma production. Five of 22 HLA-A2 and HLA-A3 volunteers displayed PfCS102 specific IFN-gamma secreting CD8(+) T cell responses. Responses were only marginally boosted after the 3(rd) vaccination and remained stable for 6 months. For both adjuvants, the dose of 10 microg was less immunogenic in comparison to 30 and 100 microg that induced similar responses. AS02A formulations with 30 microg or 100 microg PfCS102 induced about 10-folds higher antibody and IFN-gamma responses than Montanide formulations.Conclusions/significancePfCS102 peptide was safe and highly immunogenic, allowing the design of more advanced trials to test its potential for protection. Two or three immunizations with a dose of 30 microg formulated with AS02A appeared the most appropriate choice for such studies.Trial registrationSwissmedic.ch 2002 DR 1227.

Project description:Phospholipid biosynthesis plays a key role in malarial infection and is regulated by CCT (CTP:phosphocholine cytidylyltransferase). This enzyme belongs to the group of amphitropic proteins which are regulated by reversible membrane interaction. To assess the role of the putative membrane-binding domain of Plasmodium falciparum CCT (PfCCT), we synthesized three peptides, K21, V20 and K54 corresponding to residues 274-294, 308-327 and 274-327 of PfCCT respectively. Conformational behaviour of the peptides, their ability to bind to liposomes and to destabilize lipid bilayers, and their insertion properties were investigated by different biophysical techniques. The intercalation mechanisms of the peptides were refined further by using surface-pressure measurements on various monolayers at the air/water interface. In the present study, we show that the three studied peptides are able to bind to anionic and neutral phospholipids, and that they present an alpha-helical conformation upon lipid binding. Peptides V20 and the full-length K54 intercalate their hydrophobic parts into an anionic bilayer and, to a lesser extent, a neutral one for V20. Peptide K21 interacts only superficially with both types of phospholipid vesicles. Adsorption experiments performed at the air/water interface revealed that peptide K54 is strongly surface-active in the absence of lipid. Peptide V20 presents an atypical behaviour in the presence of phosphatidylserine. Whatever the initial surface pressure of a phosphatidylserine film, peptide V20 and phosphatidylserine entities seem linked together in a special organization involving electrostatic and hydrophobic interactions. We showed that PfCCT presents different lipid-dependence properties from other studied CCTs. Although the lipid-binding domain seems to be located in the C-terminal region of the enzyme, as with the mammalian counterpart, the membrane anchorage, which plays a key role in the enzyme regulation, is driven by two alpha-helices, which behave differently from one another.

Dataset Information

Synthetic Evaluation of Standard and Microwave-Assisted Solid Phase Peptide Synthesis of a Long Chimeric Peptide Derived from Four Plasmodium falciparum Proteins.

Publications

Synthetic Evaluation of Standard and Microwave-Assisted Solid Phase Peptide Synthesis of a Long Chimeric Peptide Derived from Four <i>Plasmodium falciparum</i> Proteins.

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