Biomedical subjects

Enrique A Dalmasso

Publications and source records attributed to Enrique A Dalmasso.

5 recordsLinked to original sources

PubMed2005

SELDI-TOF-MS ProteinChip array profiling of tears from patients with dry eye.

PURPOSE: Protein and peptides in tears play an important role in ocular surface diseases. In previous studies, changes have been demonstrated in the electrophoretic protein profiles of patients with dry eye. The purpose of this work was to determine the usefulness of surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) ProteinChip Array (Ciphergen Biosystems, Inc., Fremont, CA) technology for the automated analysis of proteins and peptides in tear fluid. METHODS: Patients with dry eye (DRY, n = 88) and healthy subjects (CTRL, n = 71) were examined. Their tear proteins were analyzed using SELDI-TOF-MS ProteinChip Arrays with three different chromatographic surfaces (CM10 cation exchange, Q10 anion exchange, and H50 reversed-phase) prepared by means of a laboratory liquid-handling robotic workstation. The data were analyzed by multivariate statistical techniques and artificial neural networks, and the most important biomarkers were purified and identified by tandem MS. RESULTS: Complex patterns of tear proteins and peptides were detected. The different chromatographic surfaces revealed the selective enrichment of proteins such as lipocalin and lysozyme. Discriminant analysis demonstrated highly significant changes in the protein profiles in patients with dry eye (P < 0.001). With a seven-peptide multimarker panel, an artificial neural network could differentiate between patients with dry eye and healthy individuals with a specificity and sensitivity of 90%. The identification of biomarkers revealed an increase of inflammatory markers in patients with dry eye and a decrease of some proteins that may have protective functions. CONCLUSIONS: The SELDI-TOF-MS technology seems to be ideally suitable for the mass screening of peptides and proteins in tears. This highly sensitive approach dramatically reduces the analysis time and provides protein profiles with great mass accuracy. Thus, it may become a very useful tool in the search for potential biomarkers for diagnosis and new therapeutics in ocular diseases such as dry eye.

Biomarkers↗

PubMed2004

Serum protein profile alterations in hemodialysis patients.

BACKGROUND: Serum protein profiling patterns can reflect the pathological state of a patient and therefore may be useful for clinical diagnostics. Here, we present results from a pilot study of proteomic expression patterns in hemodialysis patients designed to evaluate the range of serum proteomic alterations in this population. METHODS: Surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS) was used to analyze serum obtained from patients on periodic hemodialysis treatment and healthy controls. Serum samples from patients and controls were first fractionated into six eluants on a strong anion exchange column, followed by application to four array chemistries representing cation exchange, anion exchange, metal affinity and hydrophobic surfaces. A total of 144 SELDI-TOF-MS spectra were obtained from each serum sample. RESULTS: The overall profiles of the patient and control samples were consistent and reproducible. However, 30 well-defined protein differences were observed; 15 proteins were elevated and 15 were decreased in patients compared to controls. Serum from 1 patient exhibited novel protein peaks suggesting possible additional changes due to a secondary disease process. CONCLUSION: SELDI-TOF-MS demonstrated consistent serum protein profile differences between patients and controls. Similarity in protein profiles among dialysis patients suggests that patient physiological responses to end-stage renal disease and/or dialysis therapy have a major effect on serum protein profiles.

Adult↗

PubMed2004

Estriol bound and ligand-free structures of sterol 14alpha-demethylase.

Sterol 14alpha-demethylases (CYP51) are essential enzymes in sterol biosynthesis in eukaryotes and drug targets in antifungal therapy. Here, we report CYP51 structures in ligand-free and estriol bound forms. Using estriol as a probe, we determined orientation of the substrate in the active site, elucidated protein contacts with the invariant 3beta-hydroxy group of a sterol, and identified F78 as a key discriminator between 4alpha-methylated and 4alpha,beta-dimethylated substrates. Analysis of CYP51 dynamics revealed that the C helix undergoes helix-coil transition upon binding and dissociation of a ligand. Loss of helical structure of the C helix in the ligand-free form results in an unprecedented opening of the substrate binding site. Upon binding of estriol, the BC loop loses contacts with molecular surface and tends to adopt a closed conformation. A mechanism for azole resistance in the yeast pathogen Candida albicans associated with mutations in the ERG11 gene encoding CYP51 is suggested based on CYP51 protein dynamics.

Amino Acid Sequence↗

PubMed2002

Contribution of human alpha-defensin 1, 2, and 3 to the anti-HIV-1 activity of CD8 antiviral factor.

It has been known since 1986 that CD8 T lymphocytes from certain HIV-1-infected individuals who are immunologically stable secrete a soluble factor, termed CAF, that suppresses HIV-1 replication. However, the identity of CAF remained elusive despite an extensive search. By means of a protein-chip technology, we identified a cluster of proteins that were secreted when CD8 T cells from long-term nonprogressors with HIV-1 infection were stimulated. These proteins were identified as alpha-defensin 1, 2, and 3 on the basis of specific antibody recognition and amino acid sequencing. CAF activity was eliminated or neutralized by an antibody specific for human alpha-defensins. Synthetic and purified preparations of alpha-defensins also inhibited the replication of HIV-1 isolates in vitro. Taken together, our results indicate that alpha-defensin 1, 2, and 3 collectively account for much of the anti-HIV-1 activity of CAF that is not attributable to beta-chemokines.

Amino Acid Sequence↗

PubMed2002

Current achievements using ProteinChip Array technology.

Because of its inherent flexibility, the ProteinChip Array platform has demonstrated utility into basic research as well as clinical research. In the domain of basic research, it has been used to examine protein modifications, characterize protein-protein interactions and study signal transduction and enzymatic pathways. In clinical research, it has been used to elucidate and identify biomarkers of disease, and as a platform for predictive medicine.

Animals↗