Abel Baerga-Ortiz

University of Puerto Rico, Medical Sciences Campus · Department of Biochemistry

The development of novel antiplasmodial compounds with broad-spectrum activity against different stages of Plasmodium parasites is crucial to prevent malaria disease and parasite transmission. This study evaluated the antiplasmodial activity of seven novel hydrazone compounds (referred to as CB compounds: CB-27, CB-41, CB-50, CB-53, CB-58, CB-59, a...

We have developed a pipeline to express, purify, and characterize HIV envelope protein (Env) gp145 from Chinese hamster ovary cells, to accelerate the production of a promising vaccine candidate. First in shake flasks, then in bioreactors, we optimized the growth conditions. By adjusting the pH to 6.8, we increased expression levels to 101 mg/L in...

The Uropathogenic Specific Protein (USP) is an E. coli genotoxin nuclease primarily associated with urinary tract infections. Previous efforts in our laboratory found the USP gene in stool samples; more frequently found in stool samples from adenoma and CRC patients than in samples from healthy individuals. New efforts in our laboratory aim to find...

A fast PCR-assisted impedimetric biosensor was developed for the selective detection of the clbN gene from the polyketide synthase (pks) genomic island in real Escherichia coli samples. This genomic island is responsible for the production of colibactin, a harmful genotoxin that has been associated with colorectal cancer. The experimental protocol...

A reversed phase high performance liquid chromatography (RP-HPLC) method was developed for the quantitative determination of recombinant HIV-1 gp145 produced in CHO-K1 cells, as measured directly from culture supernatants. Samples were diluted in 50% D-PBS and 50% PowerCHO-2 (PC2) spent medium, and resolved on a Zorbax 300SB-C8 Rapid Resolution (2....

The gut microbiome has been found to impact host predisposition to disease. Previous research shows that certain species of microbes, encoding pro-inflammatory factors, increase the probability of developing dysplasia and colorectal cancer (CRC). Recently, through the oligotyping approach, we found genetic variants for an amplified region of , a ge...

The development of a prophylactic vaccine against the human immunodeficiency virus (HIV) is of paramount importance in the global drive to halt the spread of the virus. Even after the successful discovery and initial testing of a vaccine candidate, there are hurdles associated with production yield, purification strategy, and in vitro stability tha...

The envelope glycoprotein (Env) of the human immunodeficiency virus (HIV), has been the primary target for the development of a protective vaccine against infection. The extensive N-linked glycosylation on Env is an important consideration as it may affect efficacy, stability, and expression yields. The expression host has been shown to influence t...

Colibactin is a natural product made by numerous strains of E. coli that harbor the pks genomic island. The deletion of one of the genes within the pks island, the peptidase clbP, has been found to disrupt the maturation of colibactin, thus promoting the accumulation in the periplasmic space of numerous biosynthesis intermediates, some of which hav...

The envelope glycoprotein (Env) of the human immunodeficiency virus (HIV), has been the primary target for the development of a protective vaccine against infection. The extensive N -linked glycosylation on Env is an important consideration as it may affect efficacy, stability, and expression yields. The expression host has been shown to influence...

Plasmodium falciparum parasites are increasingly drug-resistant, requiring the search for novel antimalarials with distinct modes of action. Enzymes in the glutathione pathway, including glutathione S-transferase (GST), show promise as novel antimalarial targets. This study aims to better understand the biological function of Plasmodium GST, assess...

The gut microbiota has been implicated in a number of normal and disease biological processes. Recent studies have identified a subset of gut bacterial genes as potentially involved in inflammatory processes. In this work, we explore the sequence variability for some of these bacterial genes using a combination of deep sequencing and oligotyping, a...

Gut bacterial toxins are thought to contribute to the development of colorectal cancer (CRC). This study examines the presence of specific gut bacterial toxin genes in stool samples from individuals with colorectal neoplasia (adenomas and/or CRC). The presence of bacterial genes encoding genotoxic or pro-inflammatory factors (pks, tcpC, gelE, cnf-1...

Outer membrane vesicles (OMV) are proteoliposomes secreted by almost all gram‐negative bacteria with several functions like the transportation of bacterial virulence and survival factors. OMV provide an insoluble secretion pathway for gram‐negative bacteria and are responsible for the release of outer membrane and periplasm content out of the cell....

Background: The human gut microbiota is a dynamic community of microorganisms that mediate important biochemical processes. Differences in the gut microbial composition have been associated with inflammatory bowel diseases (IBD) and other intestinal disorders. In this study, we quantified and compared the frequencies of eight genotoxic and/or pro-i...

Almost all gram‐negative bacteria secrete sphere‐like proteoliposomes named outer membrane vesicles (OMVs). These vesicles provide the bacteria a vehicle for the secretion of outer membrane molecules and periplasmic content. The pks island is a gene cluster present in some strains of bacteria in the human gut microbiota, which has been correlated w...

The biosynthesis of polyunstaurated fatty acids (PUFA) in certain bacterial species, is regulated by processes that are not fully understood. One possible factor in the modulation of gene expression in PUFA biosynthesis is pfaR, a putative transcriptional regulator whose gene is encoded directly upstream of pfaA in some bacterial species but entire...

FabA and FabZ are the two dehydratase enzymes in Escherichia coli that catalyze the dehydration of acyl intermediates in the biosynthesis of fatty acids. Both enzymes form obligate dimers in which the active site contains key amino acids from both subunits. While FabA is a soluble protein that has been relatively straightforward to express and to p...

Despite increasing adherence to routine screening and advances in therapeutic strategies, CRC is the 1st and 3rd leading cause of cancer-related death in Puerto Rico and the United States (US), respectively. Currently, CRC screening is the primary means for prevention; however, 60% of CRC patients are diagnosed at more advanced, less treatable stag...

Background: Although colorectal cancer (CRC) is a potentially preventable disease, it is still one of the most common and deadliest malignancies worldwide. Despite increasing adherence to routine screening and advances in therapeutic strategies, CRC is the 1st and 3rd leading cause of cancer-related death in Puerto Rico and the United States (US),...

All gram‐negative bacteria secrete outer membrane vesicles (OMV) that have several functions, among them the transportation of bacterial virulence and survival factors. It has been found by other groups that certain OMV promote aberrant growth in mammalian cells upon contact. OMV provide an insoluble secretion pathway for gram‐negative bacteria and...

Background: Despite increasing adherence to routine screening and advances in therapeutic strategies, colorectal cancer (CRC) is still the1st and 3rd leading cause of cancer death in Puerto Rico and the US, respectively. The etiology of CRC is complex; however, environmental factors including diet, the gut microbiota, and inflammation are accepted...

Although predominantly associated with health benefits, the gut microbiota has also been shown to harbor genes that promote inflammation. In this work, we report a method for the direct detec-tion and quantification of these pro-inflammatory bacterial genes by PCR and qPCR in DNA ex-tracted from human stool samples. PCR reactions were performed to...

The enzyme telomerase is present in about 85% of human cancers which makes it not only a good target for cancer treatment but also an excellent marker for cancer detection. Using a single stranded DNA probe specific for telomerase binding and reverse transcription tethered to an interdigital gold electrode array surface, the chromosome protection p...

DNA sensors are innovative diagnostic tools that have gained broad acceptance in recent years. Different procedures have been used to develop these sensors. One of the most frequently used method is the self-assemble monolayer (SAMs) that provides advantages in terms of simplicity, efficiency and cost of electrode design. Gold surface modification...

The human intestinal microbiota plays a key role in the maintenance of health and also in the development of intestinal disorders. Numerous studies have established the definitive association between the presence of certain intestinal bacteria in the flora and the development of gastrointestinal disorders. In this work, we developed an rtPCR assay...

Polyunsaturated fatty acids are made in some strains of deep-sea bacteria by multidomain proteins that catalyze condensation, ketoreduction, dehydration and enoyl-reduction. In this work, we have employed the Udwary-Merski Algorithm sequence analysis tool to define the boundaries that enclose the dehydratase domains in a polyunsaturated fatty acid...

The polyunsaturated fatty acid (PUFA) synthases from deep-sea bacteria invariably contain multiple acyl carrier protein (ACP) domains in tandem. This conserved tandem arrangement has been implicated in both amplification of fatty acid production (additive effect) and in structural stabilization of the multidomain protein (synergistic effect). While...

Thioesterase activity is typically required for the release of products from polyketide synthase enzymes, but no such enzyme has been characterized in deep-sea bacteria associated with the production of polyunsaturated fatty acids. In this work, we have expressed and purified the Orf6 thioesterase from Photobacterium profundum. Enzyme assays reveal...

Increasing the production of fatty acids by microbial fermentation remains an important step towards the generation of biodiesel and other portable liquid fuels. In this work, we report an Escherichia coli strain engineered to overexpress a fragment consisting of four dehydratase domains from the polyunsaturated fatty acid (PUFA) synthase enzyme co...

Polyunsaturated fatty acids (PUFAs) are important components of human health and important ingredients in biodiesel preparations. PUFAs from deep‐sea bacteria are synthesized by a modular polyketide synthase, which contains several domains including two dehydratase (DH) domains responsible for the introduction of double bonds. In order to study dou...

Acyl carrier protein (ACP) is a highly conserved protein domain that plays a key role in the biosynthesis of fatty acids and polyketides. The polyunsaturated fatty acid synthase complex from deep‐sea bacteria contains a total of five ACP domains in tandem. No satisfying explanation exists for the selection of this rare arrangement. In an effort to...

Polyunsaturated fatty acids are made in deep‐sea organisms by the activity of a polyketide synthase multienzyme. The final step in the biosynthesis of polyketides and fatty acids is catalyzed by the activity of the thioesterase (TE) domain, which cleaves the final product off of the carrier protein. However, no TE domain has been identified in any...

Not a matter of preference: The polyketide borrelidin contains both a trans and a cis double bond. The corresponding dehydratases in the polyketide synthase are found to have identical preferences for thioester substrates in vitro and to generate trans double bonds, undermining the idea that cis double bonds arise from dehydration of 3S rather than...

Malaria parasites contain a complete glutathione (GSH) redox system, and several enzymes of this system are considered potential targets for antimalarial drugs. Through generation of a gamma-glutamylcysteine synthetase (gamma-GCS)-null mutant of the rodent parasite Plasmodium berghei, we previously showed that de novo GSH synthesis is not critical...

Acyl carrier protein (ACP) is a highly conserved protein domain that participates in the biosynthesis of fatty acids and polyketides. Most enzyme complexes contain either one or many of these domains arranged in a modular fashion. However, the polyketide synthase (PKS) complex responsible for the production of omega‐3 fatty acids in deep‐sea bacter...

Polyunsaturated fatty acids (PUFAs) are essential membrane components in higher eukaryotes and important components of human health. PUFAs from deep‐sea bacteria are synthesized by a modular polyketide synthase consisting of several domains including two dehydratase (DH) domains, DH1 and DH2, which are responsible for the introduction of cis or tra...

The polyketide synthase (PKS) genes responsible for eicosapentanoic acid (EPA) biosynthesis in the deep‐sea bacterium, Photobacterium profundum, have been cloned and sequenced, being a total of five polyunsaturated fatty acid (pfa) genes (pfaA, pfaB, pfaC, pfaD and Orf6) enough for EPA production. One of these enzymes is Orf6, a putative thioestera...

Marine ecosystems are a source of biologically active compounds, many of which are currently in clinical use. With the goal of increasing the availability and the chemical diversity of these important compounds, more researchers are applying the tools of biotechnology to the discovery and production of marine natural products. This review summarize...

A number of alanine and more conservative mutants of residues in the fourth domain of thrombomodulin (TM) were prepared and assayed for protein C activation and for thrombin binding. Several of the alanine mutations appeared to cause misfolding or structural defects as assessed by poor expression and/or NMR HSQC experiments, while more conservative...

The ketoreductase (KR) domains eryKR(1) and eryKR(2) from the erythromycin-producing polyketide synthase (PKS) reduce 3-ketoacyl-thioester intermediates with opposite stereospecificity. Modeling of eryKR(1) and eryKR(2) showed that conserved amino acids previously correlated with production of alternative alcohol configurations lie in the active si...

Ketoreductase (KR) activities help determine the stereochemistry of the products of modular polyketide synthases (PKSs). For example, domains eryKR(1) and eryKR(2), contained, respectively, in the first and second extension modules of the erythromycin-producing PKS, reduce 3-ketoacyl-thioester intermediates with opposite stereospecificity. Amino ac...

A system is reported for the recombinant expression of individual ketoreductase (KR) domains from modular polyketide synthases (PKSs) and scrutiny of their intrinsic specificity and stereospecificity toward surrogate diketide substrates. The eryKR(1) and the tylKR(1) domains, derived from the first extension module of the erythromycin PKS and the t...

A system is reported for the recombinant expression of individual ketoreductase (KR) domains from modular polyketide synthases (PKSs) and scrutiny of their intrinsic specificity and stereospecificity toward sur-rogate diketide substrates. The eryKR 1 and the tylKR 1 domains, derived from the first extension module of the erythromycin PKS and the ty...

This protocol describes amide proton exchange experiments that probe for changes in solvent accessibility at protein-protein interfaces. The simplest version of the protocol, termed the "on-exchange" experiment, detects protein-protein interfaces by taking advantage of the fact that solvent deuterium oxide (D2O) molecules are excluded from the surf...

Methods are presented for monitoring solvent accessibility of protein-ligand and protein-protein interfaces. The kinetics of solvent accessibility at the protein-protein interface is monitored by amide hydrogen/deuterium (H/2H) exchange detected by matrix assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). A stra...

Thrombin binds thrombomodulin (TM) at anion binding exosite 1, an allosteric site far from the thrombin active site. A monoclonal antibody (mAb) has been isolated that competes with TM for binding to thrombin. Complete binding kinetic and thermodynamic profiles for these two protein-protein interactions have been generated. Binding kinetics were me...

Mammalian fatty acid synthase is a classic example of a chain-building multienzyme. A cornerstone of its mechanism has been the obligatory collaboration of two identical subunits, with fatty acyl intermediates transferring between them. Now, fresh evidence has upset this view.

Mammalian fatty acid synthase is a classic example of a chain-building multienzyme. A cornerstone of its mechanism has been the obligatory collaboration of two identical subunits, with fatty acyl intermediates transferring between them. Now, fresh evidence has upset this view.

The epitope of a monoclonal antibody raised against human thrombin has been determined by hydrogen/deuterium exchange coupled to MALDI mass spectrometry. The antibody epitope was identified as the surface of thrombin that retained deuterium in the presence of the monoclonal antibody compared to control experiments in its absence. Covalent attachmen...

The kinetics of solvent accessibility at the protein-protein interface between thrombin and a fragment of thrombomodulin, TMEGF45, have been monitored by amide hydrogen/deuterium (H/2H) exchange detected by MALDI-TOF mass spectrometry. The interaction is rapid and reversible, requiring development of theory and experimental methods to distinguish H...

The rate constants for the binding interaction between thrombin and a fully active fragment of its anticoagulant cofactor, thrombomodulin, have been determined by surface plasmon resonance. At physiological ionic strength, the ka was 6.7 × 106 M−1 s−1 and the dissociation rate constant was 0.033 s−1. These extremely fast association and dissociatio...

Astroglial cells secrete a variety of factors that contribute to the regulation of neurite initiation and continued outgrowth, among them proteases and protease inhibitors. An alteration in the balance between these proteins has been implicated in Alzheimer's disease, resulting in an accumulation of thrombin:protease nexin 1 (PN1) complexes in the...