Marco António Gomes Saraiva
@tecnico.ulisboa.pt
RESEARCH INTERESTS
Biophysics
Scopus Publications
- The amine-imine creatinine tautomerism observed in both the solution and gas phases
Marco A. Saraiva, M. Conceição Oliveira
Journal of Molecular Structure, 2026 - Hydrophobic Hydration and Light Transport in α-Synuclein Protein Solutions in the Near-Infrared
Marco A. Saraiva
Applied Spectroscopy, 2025
Currently, there is increasing interest in identifying the mechanistic characteristics of the α-synuclein amyloid protein aggregation during its early stages. The initiation of amyloid protein incubation was investigated by applying the concepts of hydrophobic hydration in the early-formed protein aggregates and the light transport in the protein samples by using near-infrared light. These are unexplored concepts in amyloid protein aggregation research. Early-formed protein aggregates develop solvent-exposed hydrophobic residue segments, and intramolecular and intermolecular interactions can be identified by hydrophobic hydration, while consecutive intramolecular interactions can cancel this effect. In the light transport within protein samples, at low protein concentrations, the early-formed protein aggregates achieve stability, whereas at higher concentrations, such as those found in neuronal synapses (∼50 µM), the early-formed aggregates continue to develop. - Temperature-Driven Stopped-Flow Experiments for Investigating the Initial Aggregation of the α-Synuclein Amyloid Protein, Focusing on Active and Inactive Phases
Marco A. Saraiva
Journal of Fluorescence, 2025
The primary objective of this research is to further examine the events occurring during the active or burst phase by focusing on the aggregation of the Syn amyloid protein. Regarding this aspect, it was initially conducted rapid temperature variations using stopped-flow spectrometry and tyrosyl group fluorescence emission detection, within the initial 500 milliseconds in buffered Syn solutions at pH 7, exploring various temperature ranges to investigate protein aggregation. The results obtained were contrasted with results obtained for the Nα-acetyl-L-tyrosinamide (NAYA) parent compound in the same conditions. The utilization of the NAYA compound is suitable as it mimics the peptide bonds in proteins and contains a tyrosyl group resembling the four tyrosyl groups found in the Syn protein structure (the protein has no tryptophan residues). Furthermore, the NAYA compound adopts an intramolecularly hydrogen-bonded structure even in an aqueous solution, similar to the interactions seen in the hydrophilic face of β-sheets. Additionally, the Syn protein system can exhibit the presence of β-sheets as a result of the existence of very low abundant Syn amyloid precursor forms or nuclei during the initial stages of the protein aggregation. Thus, a relationship is present between the molecular processes in the NAYA and Syn protein systems, making the NAYA’s application crucial in this research. Moreover, to aid in understanding the results, it was also compared the events during the quiescent or inactive phase (30–500 milliseconds) with those in the burst phase (up to 10 milliseconds) using stopped-flow spectrometry conditions. Steady-state measurements were beneficial in comprehending the occurrences in both the quiescent and burst phases examined. Although protein aggregation and disaggregation were observed during the quiescent phase, determining these processes in the burst phase was more challenging. In the latter case, the aggregation of the Syn protein is actually initiated by the interaction of the intrinsically disordered Syn monomers. In the quiescent phase, first-order rate constants were measured and analysis showed that Syn protein aggregation and disaggregation occur simultaneously. At lower temperatures, early protein disaggregation outweighs protein aggregation whereas at higher temperatures protein disaggregation and aggregation are rather similar. It is also need to highlight that the burst phase, while distinct from the quiescent phase, can be considered as a possible structural phase for obtaining details about the aggregation of this specific disordered protein in solution on a very short timescale. - Determination of the size parameters of α-synuclein amyloid precursor forms through DLS analysis
Marco A. Saraiva
European Biophysics Journal, 2025
Currently, there is an increased interest in identifying the characteristics of amyloid aggregates in the initial stages of amyloid formation. The aggregation mechanism of the α-synuclein (Syn) amyloid protein, which has been extensively studied, is still not fully understood. I show that with conventional dynamic light scattering (DLS) technique, the measurements of the dimensions of Syn amyloid precursor forms can be done early in the protein incubation. Additionally, the early aggregation of the Syn protein was initially studied by analyzing autocorrelation functions from fit distributions up to 104 µs in the initial DLS measurements, specifically within the first 21 min. Investigation was conducted on the variation in the pH of the Syn solution throughout time. Based on DLS data, large Syn aggregated species formed from the monomer protein species. Afterward, I generated the autocorrelation functions based on the original DLS data, extending the fit distributions up to 105 µs and noticed the existence of elongated Syn amyloid precursor forms in the protein solutions. Because the length of the elongated Syn amyloid precursor forms closely matches the wavelength of the incident light, the combination of translational diffusion Dt and rotational diffusion Dr in the decay rates enabled the measurement of their geometric dimensions through DLS. The improved precision of the fitted distributions I offered resulted in a new interpretation for the Syn protein aggregation in the initial stages. Overall, the methodology used in this study could be an effective strategy for examining how Syn amyloid precursor forms develop over time. - Burst Phase Analysis of the Aggregation Prone α-synuclein Amyloid Protein
Marco A. Saraiva, M. Helena Florêncio
Journal of Fluorescence, 2024
While some studies inferred that valid information can be retrieved for the refolding of proteins and consequent identification of folding intermediates in the stopped-flow spectrometry collapse phase, other studies report that these burst phase folding intermediates can be questioned, implying a solvent-dependent modification of the still unfolded polypeptide chain. We therefore decided to investigate the burst phase occurring for the α-synuclein (Syn) amyloid protein by stopped-flow spectrometry. Solvent-dependent modification effects indeed occurred for the Nα-acetyl-L-tyrosinamide (NAYA) parent small compound and for the folded monomeric ubiquitin protein. More complex was the burst phase analysis of the disordered Syn amyloid protein. While this amyloid protein was determined to be aggregated at pH 7 and pH 2, in particular, this protein at pH 3 appears to be in a monomeric state in the burst phase analysis performed. In addition, the protein at pH 3 appears to suffer a hydrophobic collapse with the formation of a possible folded intermediate. This folded intermediate seems to result from a fast contraction of the disordered amyloid polypeptide chain, which is proceeded by an expansion of the protein, due to the occurrence of solvent-dependent modification effects in a milliseconds time scale of the burst phase. Generally, it can be argued that both literature criteria of solvent-dependent modifications of the disordered Syn amyloid protein and of the formation of its possible folded intermediate are very likely to occur in the burst phase. - Initial Effect of Temperature Rise on α-Synuclein Aggregation – Entropic Forces Drive the Exposure of Protein Hydrophobic Groups Probed by Fluorescence Spectroscopy
Marco A. Saraiva, M. Helena Florêncio
Journal of Fluorescence, 2023
The aberrant formation of α-synuclein (Syn) aggregates, varying in size, structure and morphology, has been linked to the development of Parkinson’s disease. In the early stages of Syn aggregation, large protein amyloid aggregates with sizes > 100 nm in hydrodynamic radius have been noticed. These low overall abundant large Syn aggregates are notoriously difficult to study by conventional biophysical methods. Due to the growing importance of studying the early stages of Syn aggregation, we developed a strategy to achieve this purpose, which is the study of the initial effect of the Syn protein aqueous solutions temperature rise. Therefore, the increase of the Syn aqueous solutions entropy by the initial effect of the temperature rise led to the exposure of the protein hydrophobic tyrosyl groups by not interfering with this amyloid protein aggregation. As an attempt to interpret the degree of the referred protein tyrosyl groups exposure, the classic rotameric conformations of the Nα-acetyl-L-tyrosinamide (NAYA) parent compound were used. For both NAYA and Syn, it was determined that the classic rotameric conformations involving the tyrosyl groups indeed accounted for their exposure under steady-state conditions of fluorescence, for lowest molecular species concentrations investigated at least. In this situation, Syn aggregation was observed. For the higher NAYA and Syn concentrations studied, the referred classic rotameric conformation were insufficient in such referred steady-state conditions and, for Syn, in particular, fluorescence anisotropy measurements revealed that less protein aggregation occurs along with its delay. Overall, the developed strategy by focusing on the initial effect of the temperature rise of Syn aqueous solutions in lower concentrations is suitable for informing us about the degree of this protein aggregation in solution. - Early α-synuclein aggregation is overall delayed and it can occur by a stepwise mechanism
Marco A. Saraiva, M. Helena Florêncio
Biochemical and Biophysical Research Communications, 2022 - Identification of a biological excimer involving protein–protein interactions: A case study of the α-synuclein aggregation
Marco A. Saraiva, M. Helena Florêncio
Spectrochimica Acta Part A Molecular and Biomolecular Spectroscopy, 2022 - Shear Stress Induces α-Synuclein Aggregation Due to a Less Strained Protein Backbone and Protein Tyrosyl Groups Do Not Intervene in the Aggregation
Marco A. Saraiva, Maria Helena Florêncio
Applied Sciences Switzerland, 2022
Parkinson’s disease (PD) is an increasingly prevalent and currently incurable neurodegenerative disorder. The aggregation of the amyloid disordered protein α-synuclein (Syn) has been implicated in the development of PD. In the literature, it has been suggested that tyrosine residues of Syn play an important role in the interactions established during the fibrillation process. Herein, the prevalence of the referred interactions under shear stress conditions of Nα-acetyl-L-tyrosinamide (NAYA) and of Syn solutions by using membrane centrifugal filters with different cut-off of 200 nm, 100 kDa, 50 kDa and 30 kDa, under centrifugation conditions, were investigated. In order to determine the nature of the interactions involving the protein tyrosine residues the NAYA compound, which mimics the peptide bonds in protein and also possesses a tyrosyl group similar to the tyrosyl groups found in the Syn protein molecular structure, was used. It is expected that for a small molecule, such as NAYA, no molecular association occurs, contrary to what exists in the Syn protein solutions, which can more adequately retrieve the type of interactions formed, involving the tyrosyl group. Therefore, sensing the tyrosyl group absorption, spectroscopic techniques, in particular, were used. For NAYA, an intramolecular interaction between the tyrosyl group and the peptide bond was evidenced. For NAYA and Syn, it was observed that decreasing the membrane centrifugal filters pore size, under centrifugation conditions, was concomitant with the minimization of the intramolecular interactions between the tyrosyl group and the peptide bond. With this, it is likely to assume that shear stress conditions in the Syn solutions propel protein aggregation by a less strained protein backbone. Contrary to the centrifugation of NAYA solutions, centrifuging Syn solutions revealed molecular association and a progressive exposure of protein tyrosyl groups to water. Thus, we can also infer that shear stress conditions in the Syn solutions cause the protein tyrosyl groups to not intervene in the protein aggregation. - Buffering capacity is determinant for restoring early α-synuclein aggregation
Marco A. Saraiva, M. Helena Florêncio
Biophysical Chemistry, 2022 - Evidence of the existence of micellar-like aggregates for α-synuclein
Marco A. Saraiva
International Journal of Biological Macromolecules, 2021 - Interpretation of α-synuclein UV absorption spectra in the peptide bond and the aromatic regions
Marco A. Saraiva
Journal of Photochemistry and Photobiology B Biology, 2020 - Mass spectrometric studies of the reaction of a blocked arginine with diketonic α-dicarbonyls
Marco A. Saraiva, Carlos M. Borges, M. Helena Florêncio
Amino Acids, 2016 - Earliest events in α-synuclein fibrillation probed with the fluorescence of intrinsic tyrosines
Marco A. Saraiva, Carla D. Jorge, Helena Santos, António L. Maçanita
Journal of Photochemistry and Photobiology B Biology, 2016 - Reactions of aminoguanidine with a-dicarbonyl compounds studied by electrospray ionization mass spectrometry
Marco A. Saraiva, Carlos M. Borges, M. Helena Florěncio
European Journal of Mass Spectrometry, 2012 - Behaviour of 4-(-2-hydroxyethyl)-1piperazineethanesulphonic acid under electrospray ionisation mass spectrometry conditions
Marco A. Saraiva, Carlos M. Borges, M. Helena Florêncio
European Journal of Mass Spectrometry, 2010 - Towards the control and inhibition of glycation - The role of the guanidine reaction center with aldehydic and diketonic dicarbonyls. A mass spectrometry study
Marco A. Saraiva, Carlos M. Borges, M. Helena Florêncio
Journal of Mass Spectrometry, 2006 - Non-enzymatic model glycation reactions - A comprehensive study of the reactivity of a modified arginine with aldehydic and diketonic dicarbonyl compounds by electrospray mass spectrometry
Marco A. Saraiva, Carlos M. Borges, M. Helena Florêncio
Journal of Mass Spectrometry, 2006 - Reactions of a modified lysine with aldehydic and diketonic dicarbonyl compounds: An electrospray mass spectrometry structure/activity study
Marco A. Saraiva, Carlos M. Borges, M. Helena Florêncio
Journal of Mass Spectrometry, 2006
