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1.
Mar Environ Res ; 161: 105129, 2020 Oct.
Article in English | MEDLINE | ID: mdl-32911251

ABSTRACT

Ghost fishing is a threat to many marine environments, as lost or discarded fishing gear (e.g., fishing lines, nets) continues to fish by entangling, damaging or killing various organisms. Among the benthic organisms that live on tropical reefs, the group probably most affected, due to their shape, are the branching corals. These corals provide refuge, foraging and breeding sites, especially for fishes and therefore impacts on coral structure could compromise the ecology of associated species. We tested if fishing lines entangled on the branching coral Millepora alcicornis would result in an increase in colony mortality, decrease in abundance and richness of fishes and changes in the behavior of associated reef fish. In the field, we estimated the volume of M. alcicornis colonies and its mortality percentages, and videos were recorded to evaluate abundance and richness of fish assemblages and fish behavior. Our results showed that coral mortality increased with increasing amounts of entangled fishing lines. Fish assemblages were similar in M. alcicornis colonies with or without entangled fishing lines. Nevertheless, we observed a significant decrease in the frequency of feeding attempts in two herbivore fish species (Acanthurus bahianus and Ophioblennius trinitatis) that play an important role in coral-reef dynamics, controlling algae abundances. Therefore, ghost fishing has negative impacts on shallow reef ecosystems, directly affecting branching corals and important coral-fish interactions. Management of tropical shallow reef environments should consider regulation and monitoring of coastal fisheries to ensure reef integrity.


Subject(s)
Anthozoa , Ecosystem , Animals , Coral Reefs , Fisheries , Fishes , Herbivory
2.
Genet Mol Res ; 16(2)2017 Jun 29.
Article in English | MEDLINE | ID: mdl-28671254

ABSTRACT

The recommendation of sugarcane clones depends on several factors, as the response or performance of the clones over different cuts or harvests. The clone by harvest interaction might be difficult to identify superior clones in the final stages of the sugarcane breeding program. Thus, the objective of this study was to investigate and describe the implications of the genotype by harvest interaction in the adaptability and stability of genotypes and delineation of mega-environments from a set of multi-environment trials. Fifteen clones and four checks were evaluated in eight environments. The trait TPH (tons of pol per hectare) was evaluated in two harvests (plant cane and ratoon cane) in 2010 and 2011. The joint analysis showed significance for harvest (H), environment (E), and genotype (G) effects. The interactions GxE, ExH, GxH, and ExGxH were also significant. The last three-way interaction indicated the differential response of the genotypes over environments, and that it depends on the harvests. The overall mean of the trials was 12.77 TPH. The coefficient of variation was 8.70% and the selective accuracy was 98.63%, indicating high experimental precision. The genotypes G4, G14, and G16 were statistically superior to the check varieties used; however, these genotypes did not show high stability as described by the additive main effects and multiplicative interaction method. There was a specific adaptation between the E7 and E5 environments and the G4 and G5 genotypes, respectively. In general, the grouping of the environments was inconsistent throughout the harvests, except for the E1 and E4 environments, which exhibited similarities for the different genotypes.


Subject(s)
Gene-Environment Interaction , Genotype , Plant Breeding/methods , Saccharum/genetics , Adaptation, Physiological , Environment , Genetic Variation , Quantitative Trait, Heritable , Saccharum/growth & development , Selection, Genetic
3.
Genet Mol Res ; 16(1)2017 Jan 23.
Article in English | MEDLINE | ID: mdl-28128412

ABSTRACT

Sweet sorghum has emerged as an alternative crop for ethanol yield. The breeding of this crop is performed to obtain cultivars with high ethanol yield, which necessarily requires associating favorable phenotypes for multiple traits. Therefore, the aims of this study were to investigate the association between agro-industrial traits related to ethanol yield and identify the promising genotypes considering multiple traits in sweet sorghum. For this purpose, we evaluated 45 genotypes using a 9 x 5 alpha-lattice experimental design with three replications. The traits measured were flowering time, plant height, tons of stalk per hectare, total soluble solids, tons of brix per hectare, juice extraction, total recoverable sugars, and ethanol yield. Analyses were performed after the recovery of inter-block information. The interrelation of the traits was described by genotype-by-trait biplot. For simultaneous selection, the Modified Mulamba and Mock index was used. For almost all of the agro-industrial traits, except for juice extraction, selective accuracy was above 70%. There were significant differences among genotypes for all the traits. The genotype-by-trait biplot evidenced a positive association between most of the traits related to ethanol yield, except for juice extraction, indicating the possibility of indirect selection to obtain more productive genotypes. Some genotypes proved to be promising based on the selection index, as they accumulated phenotypes favorable for the traits of interest.


Subject(s)
Quantitative Trait, Heritable , Selection, Genetic , Sorghum/physiology , Genetic Association Studies , Genotype , Phenotype , Plant Breeding
4.
Genet Mol Res ; 15(2)2016 May 20.
Article in English | MEDLINE | ID: mdl-27323058

ABSTRACT

Popcorn is widely consumed in Brazil, yet there are few breeding programs for this crop. Recurrent selection (RS) is a viable breeding alternative for popcorn; however, the gains achieved must be frequently checked. The aim of this study was to assess the effect of selection for grain type (round and pointed) after four cycles of phenotypic RS on the main agronomic traits of popcorn, to estimate the genetic gain achieved for the trait of expansion volume (EV), and to obtain estimates of phenotypic correlations for the main traits of the crop in the UFLA E and UFLA R populations. The zero, one, two, and three cycles of the UFLA E and UFLA R populations, the fourth cycle, and the controls IAC-112 and IAC-125 were used. The experiments were conducted at the experimental farm of Universidade Federal de Lavras (UFLA; Environment 1) and at the experimental area of the Genetics and Plant Breeding Sector of the Department of Biology at UFLA (Environment 2) in the 2010/11 crop season. Nine agronomic traits were evaluated, including EV and grain yield (GY). The UFLA R and UFLA E populations showed similar behavior for all evaluated traits. The type of grain did not affect the genetic gain for EV, which was 5 and 3.7% in each cycle carried out in the UFLA E and UFLA R population, respectively. Phenotypic selection carried out during recombination for EV is an effective method for increasing expression of the trait. EV and GY did not show a linear association.


Subject(s)
Callosities/genetics , Edible Grain/genetics , Quantitative Trait, Heritable , Selection, Genetic , Brazil , Breeding , Crosses, Genetic , Environment , Phenotype
5.
Genet Mol Res ; 14(4): 16392-402, 2015 Dec 09.
Article in English | MEDLINE | ID: mdl-26662435

ABSTRACT

Sweet sorghum has considerable potential for ethanol production due to its succulent stalks that contain directly fermentable sugars. Since many traits need to be considered in the selection process to breed superior cultivars for ethanol production, then correlations between the traits might be of use to help the breeder define optimal improvement strategies. The aim of this study was to investigate the association between the principal agro-industrial traits in sweet sorghum, and to evaluate the direct and indirect effects of primary and secondary traits on ethanol production per hectare. In total, 45 sweet sorghum genotypes (lineage/hybrids) were evaluated in an experiment designed in an alpha lattice 5 x 9. The data were analyzed using a mixed model approach. A detailed study of simple correlations was accomplished using path analysis. The experimental precision was high, with an accuracy above 76%. The various genotypes showed genetic variation for all agronomic and industrial traits, except stalk diameter. Some agro-industrial traits showed significant simple correlations with ethanol production, but according to the path analysis, some of these traits did not show a significant direct or indirect effect on ethanol production. The results highlighted the primary and secondary traits with practical relevance to sweet sorghum breeding, since they showed director indirect effects on ethanol production.


Subject(s)
Genetic Association Studies , Genotype , Quantitative Trait, Heritable , Sorghum/genetics , Sorghum/metabolism , Environment , Ethanol/metabolism
6.
Genet Mol Res ; 14(4): 14043-9, 2015 Oct 30.
Article in English | MEDLINE | ID: mdl-26535718

ABSTRACT

Genetic diversity among local accessions and varieties subsidize plant breeding programs, allowing the utilization of existing variability in plants that have already adapted to local climate conditions. An alternative to studying genetic variability is the study of diversity. The aim of this research was to study genetic diversity among sugarcane accessions and varieties used for the production of craft-distilled cachaça (distilled sugarcane alcohol) in the region of Lavras, Minas Gerais, Brazil. Using a one-way design, an experiment was conducted in the municipality of Perdões, Minas Gerais to evaluate 35 regional accessions derived from germplasm collection expeditions and four varieties. Using morphological descriptions of 46 multicategorical sugarcane characteristics, dissimilarity and Tocher cluster method analyses were performed. Based on the results, it was concluded that genetic diversity exists among the accessions evaluated for the target traits.


Subject(s)
Saccharum/genetics , Brazil , Cluster Analysis , Genetic Drift , Genetic Variation , Phenotype , Plant Breeding , Seed Bank
7.
Genet Mol Res ; 14(3): 11211-21, 2015 Sep 22.
Article in English | MEDLINE | ID: mdl-26400352

ABSTRACT

Sweet sorghum has considerable potential for ethanol and energy production. The crop is adaptable and can be grown under a wide range of cultivation conditions in marginal areas; however, studies of phenotypic stability are lacking under tropical conditions. Various methods can be used to assess the stability of the crop. Some of these methods generate the same basic information, whereas others provide additional information on genotype x environment (G x E) interactions and/or a description of the genotypes and environments. In this study, we evaluated the complementarity of two methods, GGEBiplot and Toler, with the aim of achieving more detailed information on G x E interactions and their implications for selection of sweet sorghum genotypes. We used data from 25 sorghum genotypes grown in different environments and evaluated the following traits: flowering (FLOW), green mass yield (GMY), total soluble solids (TSS), and tons of Brix per hectare (TBH). Significant G x E interactions were found for all traits. The most stable genotypes identified with the GGEBiplot method were CMSXS643 for FLOW, CMSXS644 and CMSXS647 for GMY, CMSXS646 and CMSXS637 for TSS, and BRS511 and CMSXSS647 for TBH. Especially for TBH, the genotype BRS511 was classified as doubly desirable by the Toler method; however, unlike the result of the GGEBiplot method, the genotype CMSXS647 was also found to be doubly undesirable. The two analytical methods were complementary and enabled a more reliable identification of adapted and stable genotypes.


Subject(s)
Sorghum/genetics , Adaptation, Physiological , Genome, Plant , Genomic Instability , Genotype , Plant Breeding , Sorghum/growth & development
8.
Int J Obes (Lond) ; 39(6): 977-85, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25771927

ABSTRACT

BACKGROUND/OBJECTIVES: Lymphocytes have a critical role in visceral adipose tissue (AT) inflammation. The CD28 costimulatory molecule is required for lymphocyte activation and for the development of a functional regulatory T cells (Tregs) compartment; however, its role during obesity is unknown. METHODS: During diet-induced obesity, we investigated the effects of selective interference with CD28 signaling using knockout mice (Cd28KO) and a CTLA4-Ig fusion protein inhibiting CD28-B7 interactions. RESULTS: Cd28 deficiency decreased pathogenic T cells and Treg content within AT without changing the macrophages number. Cd28KO epididymal but not subcutaneous fat was characterized by enlarged adipocytes, reduced levels of inflammatory cytokines and increased Glut4, adiponectin and lipogenic enzyme mRNA levels. This was associated with reduced inflammation, fat accumulation and enhanced glucose metabolism in liver. Weight gain and fasting glucose tolerance were not affected. CTLA4-Ig injections reduced the number of T cells in epididymal AT (epiAT) but not the inflammatory cytokines levels and failed to improve liver fat accumulation. CONCLUSIONS: Deletion of CD28 creates a new pro/anti-inflammatory balance in epiAT and liver and exerts a protective effect against hepatic steatosis.


Subject(s)
Adipose Tissue/pathology , CD28 Antigens/genetics , Fatty Liver/pathology , Gene Deletion , Inflammation/pathology , Liver/pathology , Obesity/pathology , Animals , Disease Models, Animal , Inflammation/metabolism , Insulin Resistance , Intracellular Signaling Peptides and Proteins/metabolism , Lipid Metabolism , Liver/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Tumor Necrosis Factor Receptor Superfamily, Member 7
10.
Biol Sport ; 31(1): 3-7, 2014 Mar.
Article in English | MEDLINE | ID: mdl-24917683

ABSTRACT

The aim of the present study was to investigate changes in muscle soreness, blood muscle damage markers, muscle strength and agility following an official basketball match. Eleven elite female professional basketball players (27.4 ± 4.8 years, 179.5 ± 5.5 cm, 72.0 ± 7.8 kg) of a team participated in this study. The official match was the seventh match of the season in the first phase of the Brazilian National Female Basketball Championship. Muscle soreness, plasma creatine kinase activity (CK), and myoglobin concentration (Mb) were determined before and after the match (post-match, 24 and 48 hours after the match). The 1RM strength for bench press and leg press, and the agility T test were assessed before and at 24 and 48 hours after the match. Significant increases in muscle soreness, CK and Mb were observed at 24 and 48 hours post-match (p<0.05). No significant changes in the 1RM strength and T test were detected during recovery (24 and 48 hours after the match). These results suggest that a basketball match induced limited muscle damage with minimal effect on performance during recovery. The small increase in muscle damage markers following a basketball match did not affect strength and agility performance.

11.
J Sports Med Phys Fitness ; 51(4): 676-82, 2011 Dec.
Article in English | MEDLINE | ID: mdl-22212272

ABSTRACT

AIM: The aim of this paper was to examine the effects of resistance training periodization on the performance and salivary hormone-immune responses of elite female basketball players. METHODS: Twelve female athletes were monitored across a 50 day period of resistance training that emphasized strength, endurance and power. One repetition maximum (1RM) strength, maximal repetitions at 50% 1RM and vertical jump performance was assessed pre- and post-training. Saliva samples were also collected at 0700, 0930, 1100 and 1730 hours and analyzed for testosterone (T), cortisol (C) and immunoglobulin A (IgA). RESULTS: Improvements in 1RM strength, maximal repetitions and vertical jump performance were identified post-training (P<0.05). Training had no effect on salivary T and C concentrations, but the T:C ratio increased at 0730 hours (P<0.05) and IgA concentrations were lowered at 0930 and 1100 hours (P<0.05). The changes (∆ Pre-Post training) in strength and T concentrations were positively correlated at 0730 hours (P<0.05). CONCLUSION: A periodized approach to resistance training increased muscle performance in elite female basketball players, but only minor changes in the salivary T:C ratio and IgA were noted. Correlational analysis identified a possible role for early morning changes in T as a regulator of individual strength changes.


Subject(s)
Athletic Performance/physiology , Basketball/physiology , Resistance Training/methods , Saliva/metabolism , Adult , Female , Humans , Hydrocortisone/metabolism , Immunoglobulin A/metabolism , Muscle Strength , Muscle, Skeletal/physiology , Testosterone/metabolism , Time Factors , Young Adult
13.
Gene Ther ; 15(12): 942-52, 2008 Jun.
Article in English | MEDLINE | ID: mdl-18323795

ABSTRACT

Further development of haematopoietic stem cell (HSC) gene therapy will depend on enhancement of gene transfer safety: ad hoc improvement of vector design relating to each particular disease is thus a crucial issue for HSC gene therapy. We modified a previously described lentiviral vector by adding the Emumar B-specific enhancer to a human CD19 promoter-derived sequence (Mol Ther 2004;10:45-56). We thus significantly improved the level of expression of the green fluorescent protein (GFP) reporter gene while retaining the specificity of expression in B-cell progeny of transduced human CD34+ progenitor cells obtained from cord blood or adult bone marrow. Indeed, GFP was strongly expressed from early medullary pro-B cells to splenic mature B cells whereas transgene expression remained low in transduced immature progenitors as in myeloid and T-lymphoid progeny retrieved from xenografted NOD/SCID/gammac(null) mice. Using this lentiviral vector, we further demonstrated the possibility to express a functional human BTK protein in long-term human CD34+ cell B-lymphoid progeny. This newly designed lentiviral vector fulfils one of the pre-requisites for the development of efficient and safe gene therapy for X-linked agammaglobulinaemia, the most common primary humoral immunodeficiency disorder.


Subject(s)
Agammaglobulinemia/therapy , B-Lymphocytes/metabolism , Genetic Therapy/methods , Lentivirus/genetics , Protein-Tyrosine Kinases/genetics , Agammaglobulinaemia Tyrosine Kinase , Agammaglobulinemia/immunology , Agammaglobulinemia/metabolism , Animals , Antigens, CD34/immunology , B-Lymphocytes/immunology , Cell Line , Cells, Cultured , Gene Expression , Genetic Engineering , Genetic Vectors , Green Fluorescent Proteins/genetics , Hematopoietic Stem Cells/immunology , Hematopoietic Stem Cells/metabolism , Humans , Mice , Mice, SCID , Models, Animal , Transduction, Genetic/methods , Transgenes , Transplantation, Heterologous
14.
J Exp Bot ; 57(4): 971-83, 2006.
Article in English | MEDLINE | ID: mdl-16488915

ABSTRACT

The RAD21/REC8 gene family has been implicated in sister chromatid cohesion and DNA repair in several organisms. Unlike most eukaryotes, Arabidopsis thaliana has three RAD21 gene homologues, and their cloning and characterization are reported here. All three genes, AtRAD21.1, AtRAD21.2, and AtRAD21.3, are expressed in tissues rich in cells undergoing cell division, and AtRAD21.3 shows the highest relative level of expression. An increase in steady-state levels of AtRAD21.1 transcript was also observed, specifically after the induction of DNA damage. Phenotypic analysis of the atrad21.1 and atrad21.3 mutants revealed that neither of the single mutants was lethal, probably due to the redundancy in function of the AtRAD21 genes. However, AtRAD21.1 plays a critical role in recovery from DNA damage during seed imbibition, prior to germination, as atrad21.1 mutant seeds are hypersensitive to radiation damage.


Subject(s)
Arabidopsis Proteins/physiology , Arabidopsis/genetics , Arabidopsis/radiation effects , Nuclear Proteins/physiology , Arabidopsis/physiology , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Chromosomal Proteins, Non-Histone/genetics , Chromosomal Proteins, Non-Histone/metabolism , Chromosomal Proteins, Non-Histone/physiology , Cloning, Molecular , DNA Damage , Flowers/anatomy & histology , Flowers/physiology , Flowers/radiation effects , Gene Expression Regulation, Plant , Genes, Plant , Genes, Reporter , Molecular Sequence Data , Mutation , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Phenotype , RNA, Messenger/metabolism , Radiation, Ionizing , Seedlings/anatomy & histology , Seedlings/physiology , Seedlings/radiation effects , Seeds/anatomy & histology , Seeds/physiology , Seeds/radiation effects , Sequence Analysis, Protein , Sequence Homology, Nucleic Acid
15.
Genes Immun ; 4(1): 40-5, 2003 Jan.
Article in English | MEDLINE | ID: mdl-12595900

ABSTRACT

Dok proteins are adapter proteins involved in signal transduction. Several intracellular proteins expressed in lymphocytes meet the criteria of membrane-associated adapter proteins such as members of the Dok family. To understand the role and the formation of multiprotein networks involving Dok proteins in T lymphocytes, we search for potential additional members of this family. Here, we describe the two new human dok-related genes DOK4 and DOK5 and present data showing the expression of DOK4 and DOK5 genes in T cells. These genes are the orthologues of mouse Dok4 and Dok5 genes. Based on analysis of phylogenetic trees and exon/intron structure of Dok family members, DOK4 and DOK5 define a subfamily within dok genes distinct from DOK1, DOK2 and DOK3. So, Dok-4 and Dok-5 molecules constitute a new group of adapter proteins in T cells, requiring further functional analysis.


Subject(s)
Carrier Proteins/biosynthesis , Carrier Proteins/genetics , Intracellular Signaling Peptides and Proteins , Phosphoproteins , T-Lymphocytes/metabolism , Adaptor Proteins, Signal Transducing , Gene Expression Regulation/physiology , Humans , Molecular Sequence Data , Phylogeny
16.
Mol Cell Biol ; 21(14): 4515-27, 2001 Jul.
Article in English | MEDLINE | ID: mdl-11416131

ABSTRACT

The interaction of interleukin-2 (IL-2) with its receptor (IL-2R) critically regulates the T-cell immune response, and the alpha chain CD25/IL-2Ralpha is required for the formation of the high-affinity receptor. Tissue-specific, inducible expression of the IL-2Ralpha gene is regulated by at least three positive regulatory regions (PRRI, PRRII, and PRRIII), but none responded to CD28 engagement in gene reporter assays although CD28 costimulation strongly amplifies IL-2Ralpha gene transcription. By DNase I hypersensitivity analysis, we have identified a novel TCR-CD3- and CD28-responsive enhancer (CD28rE) located 8.5 kb 5' of the IL-2Ralpha gene. PRRIV/CD28rE contains a functional CRE/TRE element required for CD28 signaling. The T-cell-specific, CD28-responsive expression of the IL-2Ralpha gene appears controlled through PRRIV/CD28rE by cooperation of CREB/ATF and AP-1 family transcription factors.


Subject(s)
Blood Proteins/metabolism , CD28 Antigens/metabolism , Cyclic AMP Response Element-Binding Protein/metabolism , Enhancer Elements, Genetic , Receptors, Interleukin-2/genetics , Transcription Factor AP-1/metabolism , Transcription Factors/metabolism , Transcriptional Activation , Activating Transcription Factors , Animals , Base Sequence , COS Cells , Chlorocebus aethiops , DNA, Complementary , Humans , Jurkat Cells , Molecular Sequence Data , Receptor-CD3 Complex, Antigen, T-Cell/metabolism , Regulatory Sequences, Nucleic Acid
17.
Interv Neuroradiol ; 7(2): 121-6, 2001 Jun 30.
Article in English | MEDLINE | ID: mdl-20663337

ABSTRACT

SUMMARY: We describe a rare case of multiple arteriovenous fistulae of the spinal cord (SCAVF) in the same myelomer in a five-year-old boy. This case report consists of a trifocal SCAVF at the Th12 myelomeric level without communication between the three different fistulae. Two AVF were located posteriorly, bilateraly, in the spinal cord, fed by left and right posterior radiculopial arteries and one anteriorly in the anterior spinal axis. The venous drainage was independent for each lesion. The patient presents associated lesions characterized by cutaneous stain and inferior limb asymmetry. A metameric distribution is the explanation for the multiplicity of these lesions in a syndromic association related to Cobb syndrome. The patient was treated by transarterial embolization using glue with occlusion of the three different fistulae. The patient achieved a good improvement in neurological status.

18.
Int Immunol ; 12(11): 1547-52, 2000 Nov.
Article in English | MEDLINE | ID: mdl-11058574

ABSTRACT

The Tec protein kinase family includes Btk, Itk, Tec, Rlk and Bmx, which are critically involved in signals mediated by various cytokines and antigen receptors. Btk mutations cause severe immunodeficiencies, with defective B cell function. In T cells, Tec regulates cytokine production. However, the downstream targets of these Tec kinases are poorly defined. Here we report that overexpression of Tec in T cells can regulate gene transcription through the nuclear factor of activated T cells (NF-AT). Using different reporter gene constructs, we establish that Tec in transfected T cells dramatically induced NF-AT-dependent gene transcription, which was prevented by a dominant-negative mutant of NF-AT or by the immunosuppressive drug cyclosporin A. Tec appears to regulate NF-AT nuclear import. In addition, Tec influences cytoplasmic free calcium increase. Taken together, our results identify NF-AT as a major downstream target of Tec kinases that is critically involved in transcriptional gene regulation. These observations highlight signaling pathways regulated by Tec kinases and provide new pharmacological targets to regulate immune functions.


Subject(s)
DNA-Binding Proteins/physiology , Lymphocyte Activation , Protein-Tyrosine Kinases/physiology , Signal Transduction/immunology , T-Lymphocytes/enzymology , T-Lymphocytes/immunology , Transcription Factors/physiology , Biological Transport/immunology , Cell Line , Humans , Jurkat Cells , Mast Cells , NFATC Transcription Factors , Nuclear Proteins/physiology , Tumor Cells, Cultured
20.
J Mol Biol ; 298(1): 123-33, 2000 Apr 21.
Article in English | MEDLINE | ID: mdl-10756109

ABSTRACT

Cyclosporin A (CsA) is a potent anti-malarial compound in vitro and in vivo in mice though better known for its immunosuppressive properties in humans. Crystal structures of wild-type and a double mutant Plasmodium falciparum cyclophilin (PfCyP19 and mPfCyP19) complexed with CsA have been determined using diffraction terms to a resolution of 2.1 A (1 A=0.1 nm). The wild-type has a single PfCyP19/CsA complex per asymmetric unit in space group P1 and refined to an R-work of 0.15 and R-free of 0.19. An altered cyclophilin, with two accidental mutations, Phe120 to Leu in the CsA binding pocket and Leu171 to Trp at the C terminus, presents two complexes per asymmetric unit in the orthorhombic space group P2(1)2(1)2. This refined to an R-work of 0.18 and R-free 0.21. The mutations were identified from the crystallographic analysis and the C-terminal alteration helps to explain the different crystal forms obtained. PfCyP19 shares approximately 61 % sequence identity with human cyclophilin A (hCyPA) and the structures are similar, consisting of an eight-stranded antiparallel beta-barrel core capped by two alpha-helices. The fold creates a hydrophobic active-site, the floor of which is formed by side-chains of residues from four antiparallel beta-strands and the walls from loops and turns. We identified C-H.O hydrogen bonds between the drug and protein that may be an important feature of cyclophilins and suggest a general mode of interaction between hydrophobic molecules. Comparisons with cyclophilin-dipeptide complexes suggests that a specific C-H.O hydrogen bonding interaction may contribute to ligand binding. Residues Ser106, His99 and Asp130, located close to the active site and conserved in most cyclophilins, are arranged in a manner reminiscent of a serine protease catalytic triad. A Ser106Ala mutant was engineered to test the hypothesis that this triad contributes to CyP function. Mutant and wild-type enzymes were found to have similar catalytic properties.


Subject(s)
Antimalarials/metabolism , Cyclosporine/metabolism , Peptidylprolyl Isomerase/chemistry , Peptidylprolyl Isomerase/metabolism , Plasmodium falciparum/chemistry , Amino Acid Sequence , Amino Acid Substitution , Animals , Antimalarials/chemistry , Binding Sites , Catalysis , Conserved Sequence , Crystallography, X-Ray , Cyclosporine/chemistry , Humans , Hydrogen Bonding , Models, Molecular , Molecular Sequence Data , Mutation/genetics , Peptidylprolyl Isomerase/genetics , Plasmodium falciparum/genetics , Protein Structure, Secondary , Sequence Alignment , Structure-Activity Relationship
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