Ligand-induced changes in the binding sites of proteins

Classical molecular interaction potentials, in conjunction with other theoretical techniques, are used to analyze the dependence of the binding sites of representative proteins on the bound ligand. It is found that the ligand bound introduces in general small structural perturbations at the binding site of the protein. However, such small structural changes can lead to important alterations in the recognition pattern of the protein. The impact of these findings in docking procedures is discussed.     

A simple and rapid PCR-based method to isolate complete small macronuclear minichromosomes from hypotrich ciliates: 5S rDNA and S26 ribosomal protein gene of Oxytricha (Sterkiella) nova

Hypotrich ciliates present a macronuclear genome consisting of gene-sized instead of chromosome-sized DNA molecules. Exploiting this unique eukaryotic genome feature, we introduce, for the first time in ciliates, a rapid and easy PCR method using telomeric primers to isolate small complete macronuclear DNA molecules or minichromosomes. Two presumably abundant macronuclear DNA molecules, containing ribosomal genes, were amplified from the Oxytricha (Sterkiella) nova complete genome after using this method, and then were cloned and sequenced. The 5S rDNA sequence of O. (S.) nova is the third one reported among hypotrich ciliates; its primary and secondary structure is compared with other eukaryotic 5S rRNAs. The ribosomal protein S26 gene is the first one reported among ciliates. This "End-End-PCR" method might be useful to obtain similar gene-sized macronuclear molecules from other hypotrich ciliates, and, therefore, to increase our knowledge on ribosomal genes in these eukaryotic microorganisms.     

Sperm coating mechanism from the 1.8 A crystal structure of PDC-109-phosphorylcholine complex

Bovine seminal plasma PDC-109 binds to sperm surface choline lipids and promotes sperm capacitation by stimulating the efflux of cholesterol and phospholipids. The structure of PDC-109 with bound phosphorylcholine was solved using MAD data of a single platinum site. Its two globular (40 x 50 x 20 A(3)) Fn2 domains are linked and clustered by a short polypeptide. The choline binding sites lie at the same face of the molecule. Phosphorylcholine binds to the Fn2 domains through a cation-pi interaction between the quaternary ammonium group and a core tryptophan, plus hydrogen bonding between hydroxyls of exposed tyrosines and the phosphate group. The structure of the PDC-109-oPC complex provides a structural ground for the sperm membrane-coating mechanism underlying PDC-109-induced capacitation.     

Guanidinium and aminoimidazolinium derivatives of N-(4-piperidyl)propanamides as potential ligands for mu opioid and I2-imidazoline receptors: synthesis and pharmacological screening

Derivatives of N-(1-phenethyl-4-piperidyl)propanamides incorporating guanidinium and 2-aminoimidazolinium groups have been prepared by a synthetic approach involving first introduction of a spacer between the piperidine and the functional group by reductive amination of piperidinone. The formation of each of these functional groups was carried out using N-N'-di(tert-butoxycarbonyl)thiourea and 2-methylthioimidazolinium iodide, respectively. These structures have been designed to incorporate two pharmacologic goals into one entity. Radioligand binding assays have been used to study their affinity for opioid (mu, delta and kappa) and I2-imidazoline receptors. Two of them, 10 and 16, showed high affinity for mu opioid receptors and functionally they had moderate analgesic properties in the hot plate and writhing tests. The in vitro studies on guinea pig ileum (GPI) indicated that both compounds are mu opioid agonists. In what concerns I2-imidazoline receptor activity, these derivatives showed low affinity around 6 to 7 times less than idazoxan.     

Myxobolus desaequalis n. sp. (Myxozoa,Myxosporea), parasite of the Amazonian freshwater fish,Apteronotus albifrons (Teleostei,Apteronotidae)

Myxobolus desaequalis n. sp. is described from the gill lamellae of the freshwater fish Apteronotus albifrons, collected in the Amazon River, near the city of Salvaterra, Brazil. Large spherical plasmodia filled with disporic pansporoblasts and spores were observed. Ellipsoidal to pyriform spores are 18.3 microm length x 11.2 microm width x 4.4 microm thickness. The anterior end of the spores contain two extremely unequal pyriform polar capsules measuring: (larger): 11.2 microm length, 4.9 microm width, and an isofilar polar filament with 11 to 12 turns obliquely to the longitudinal axis; (smaller): 4.6 microm length, 2.8 microm width, and an isofilar polar filament with 4 to 5 turns, obliquely to the longitudinal axis.     

Release of NO by a nitrosyl complex upon activation by the mitochondrial reducing power

The reaction of trans-[Ru(NH(3))(4)P(OEt)(3)NO](3+) and mitochondria was investigated through differential pulse polarography and fluorimetry. The nitrosyl complex undergoes one-electron reduction centered on the NO ligand site. The reaction between the mitochondrial reductor and trans-[Ru(NH(3))(4)P(OEt)(3)NO](3+) exhibits a second order specific rate constant calculated as k=2 x 10(1) M(-1) s(-1). The reduced species, trans-[Ru(NH(3))(4)P(OEt)(3)NO](2+), quickly releases NO, yielding trans-[Ru(NH(3))(4)P(OEt)(3)H(2)O](2+). The low toxicities of both trans-[Ru(NH(3))(4)P(OEt)(3)(NO)](2+) and trans-[Ru(NH(3))(4)P(OEt)(3)H(2)O](2+) and its ability to release NO after reductive activation in a biological medium make the nitrosyl compound a useful model of a hypotensive drug.     

Steady-state luminescence investigation of the binding of Eu(III) and Tb(III) ions with synthetic peptides derived from plant thionins

This work reports Eu(III) and Tb(III) luminescence titrations in which the lanthanide ions were used as spectroscopic probes for Ca(II) ions to determine the metal binding ability of Ac-NESVKEEGGW-NH(2) and Ac-NESVKEDGGW-NH(2). These decapeptides correspond to the putative calcium binding region of the plant antifungal proteins SI-alpha1 from Sorghum bicolor and of Zeathionin from Zea mays, respectively. The luminescence spectra for the Eu(III)-decapeptide system (red emission) with the excitation at the Trp band at 280 nm showed an enhancement of the intensities of the 5D(0)-->7F(J) transitions (where J=0-4) with increments of Eu(III) ion concentration. The photoluminescence titration data of the terbium ion (green emission) in the decapeptide solutions showed intensification of the 5D(4)-->7F(J) transitions (J=0-6), similar to that observed for the Eu(III) ion. Thus, energy transfer from Ac-NESVKEEGGW-NH(2) and Ac-NESVKEDGGW-NH(2) to the trivalent lanthanide ions revealed that these peptides are capable of binding to these metal ions with association constants of the order of 10(5) M(-1). The amino acid derivative Ac-Trp-OEt also transferred energy to Tb(III) and Eu(III) ions as judged from the quenching of tryptophan luminescence. However, the energy transfers were significantly lower. Taken together the luminescence titration data indicated that Ac-NESVKEEGGW-NH(2) and Ac-NESVKEDGGW-NH(2) bind efficiently to both trivalent lanthanide ions and that these ions may be used as probes to distinguish an anionic peptide from a neutral amino acid derivative.