Structural insights into the catalytic mechanism of sphingomyelinases D and evolutionary relationship to glycerophosphodiester phosphodiesterases

Spider venom sphingomyelinases D catalyze the hydrolysis of sphingomyelin via an Mg2+ ion-dependent acid-base catalytic mechanism which involves two histidines. In the crystal structure of the sulfate free enzyme determined at 1.85 angstrom resolution, the metal ion is tetrahedrally coordinated instead of the trigonal-bipyramidal coordination observed in the sulfate bound form. The observed hyperpolarized state of His47 requires a revision of the previously suggested catalytic mechanism. Molecular modeling indicates that the fundamental structural features important for catalysis are fully conserved in both classes of SMases D and that the Class II SMases D contain an additional intra-chain disulphide bridge (Cys53-Cys201). Structural analysis suggests that the highly homologous enzyme from Loxosceles bonetti is unable to hydrolyze sphingomyelin due to the 95G1y -> Asn and 134Pro -> Glu mutations that modify the local charge and hydrophobicity of the interfacial face. Structural and sequence comparisons confirm the evolutionary relationship between sphingomyelinases D and the glicerophosphodiester phosphoesterases which utilize a similar catalytic mechanism. (c) 2006 Elsevier B.V. All rights reserved.

Structural basis for metal ion coordination and the catalytic mechanism of sphingomyelinases D

Sphingomyelinases D (SMases D) from Loxosceles spider venom are the principal toxins responsible for the manifestation of dermonecrosis, intravascular hemolysis, and acute renal failure, which can result in death. These enzymes catalyze the hydrolysis of sphingomyelin, resulting in the formation of ceramide 1-phosphate and choline or the hydrolysis of lysophosphatidyl choline, generating the lipid mediator lysophosphatidic acid. This report represents the first crystal structure of a member of the sphingomyelinase D family from Loxosceles laeta (SMase I), which has been determined at 1.75-angstrom resolution using the quick cryo-soaking technique and phases obtained from a single iodine derivative and data collected from a conventional rotating anode x-ray source. SMase I folds as an (alpha/beta)(8) barrel, the interfacial and catalytic sites encompass hydrophobic loops and a negatively charged surface. Substrate binding and/or the transition state are stabilized by a Mg2+ ion, which is coordinated by Glu(32), Asp(34), Asp(91), and solvent molecules. In the proposed acid base catalytic mechanism, His(12) and His(47) play key roles and are supported by a network of hydrogen bonds between Asp(34), Asp(52), Trp(230), Asp(233), and Asn(252).

Structure of a novel class II phospholipase D: Catalytic cleft is modified by a disulphide bridge

Phospholipases D (PLDs) are principally responsible for the local and systemic effects of Loxosceles envenomation including dermonecrosis and hemolysis. Despite their clinical relevance in loxoscelism, to date, only the SMase I from Loxosceles laeta, a class I member, has been structurally characterized. The crystal structure of a class II member from Loxosceles intermedia venom has been determined at 1.7. Å resolution. Structural comparison to the class I member showed that the presence of an additional disulphide bridge which links the catalytic loop to the flexible loop significantly changes the volume and shape of the catalytic cleft. An examination of the crystal structures of PLD homologues in the presence of low molecular weight compounds at their active sites suggests the existence of a ligand-dependent rotamer conformation of the highly conserved residue Trp230 (equivalent to Trp192 in the glycerophosphodiester phosphodiesterase from Thermus thermophofilus, PDB code: 1VD6) indicating its role in substrate binding in both enzymes. Sequence and structural analyses suggest that the reduced sphingomyelinase activity observed in some class IIb PLDs is probably due to point mutations which lead to a different substrate preference. © 2011 Elsevier Inc.

Citogenética de 13 espécies de aranhas haploginas pertencentes às famílias Pholcidae, Sicariidae e Scytodidae (Araneomorphae): evolução cromossômica, sistema cromossômico de determinação sexual e citotaxonomia

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Recent advances in the understanding of brown spider venoms: From the biology of spiders to the molecular mechanisms of toxins

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Integrated pest management programme in hospital environment

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Structural insights into substrate binding of brown spider venom class II phospholipases D

Phospholipases D (PLDs), the major dermonecrotic factors from brown spider venoms, trigger a range of biological reactions both in vitro and in vivo. Despite their clinical relevance in loxoscelism, structural data is restricted to the apo-form of these enzymes, which has been instrumental in understanding the functional differences between the class I and II spider PLDs. The crystal structures of the native class II PLD from Loxosceles intermedia complexed with myo-inositol 1-phosphate and the inactive mutant H12A complexed with fatty acids indicate the existence of a strong ligand-dependent conformation change of the highly conserved aromatic residues, Tyr 223 and Trp225 indicating their roles in substrate binding. These results provided insights into the structural determinants for substrate recognition and binding by class II PLDs.

Tratamento cirúrgico da úlcera loxoscélica: solução prática para um problema de difícil resolução

Contexto: O manuscrito descreve a reparação cirúrgica de uma úlcera crônica resultante da picada de uma aranhamarrom, um problema de difícil resolução, o qual habitualmente demora meses até a cicatrização. Descrição do caso: A vítima foi picada na coxa direita e desenvolveu fases de envenenamento típicas da picada de Loxosceles, incluindo a placa mármore, a escara e a úlcera profunda e de bordos emoldurados, características do acidente. Devido ao processo cicatricial lento, cuidados intensivos com curativos, dificuldade na deambulação, além do fator estético, optou-se por cirurgia excisional da lesão e sutura direta das bordas. O tratamento cirúrgico proporcionou cicatrização total da lesão dentro de 15 dias. Conclusões: A úlcera loxoscélica apresenta difícil cicatrização, evoluindo para a cronicidade. A cirurgia com excisão completa e síntese primária constitui técnica adequada para a cura da doença.

Estudos computacionais de esfingomielinases D: docking, dinâmica molecular e métodos híbridos QM/MM

Pós-graduação em Biofísica Molecular - IBILCE

Estudos computacionais de esfingomielinases D: docking, dinâmica molecular e métodos híbridos QM/MM

Pós-graduação em Biofísica Molecular - IBILCE

Venom composition and strategies in spiders: is everything possible?

This review on all spider venom components known by the end of 2010 bases on 1618 records for venom compounds from 174 spider species (= 0.41% of all known species) belonging to 32 families (= 29% of all existing spider families). Spiders investigated for venom research are either big (many mygalomorph species, Nephilidae, Ctenidae and Sparassidae) or medically important for humans (e.g. Loxosceles or Latrodectus species). Venom research widely ignored so far the two most species-rich families (Salticidae and Linyphiidae) and strongly neglected several other very abundant families (Araneidae, Lycosidae, Theridiidae, Thomisidae and Gnaphosidae). We grouped the known 1618 records for venom compounds into six categories: low molecular mass compounds (16 % of all compounds), acylpolyamines (11 %), linear peptides (6 %), cysteine-knotted mini-proteins (60 %), neurotoxic proteins (1 %) and enzymes (6 %). Low molecular mass compounds are known from many spider families and contain organic acids, nucleosides, nucleotides, amino acids, amines, polyamines, and some further substances, many of them acting as neurotransmitters. Acylpolyamines contain amino acids (Araneidae and Nephilidae) or not (several other families) and show a very high diversity within one species. Linear peptides, also called cytolytic, membranolytic or antimicrobial, exert a highly specific structure and are so far only known from Ctenidae, Lycosidae, Oxyopidae and Zodariidae. Cysteine-knotted mini-proteins represent the majority of venom compounds because research so far focused on them. They probably occur in most but not all spider families. Neurotoxic proteins so far are only known from theridiid spiders. Enzymes had been neglected for some time but meanwhile it becomes obvious that they play an important role in spider venoms. Sixteen enzymes either cleave polymers in the extracellular matrix or target phospholipids and related compounds in membranes. The overall structure of these compounds is given and the function, as far as it is known, is described. Since several of these component groups are presented in one average spider venom, we discuss the known interactions and synergisms and give reasons for such a functional redundancy. We also discuss main evolutionary pathways for spider venom compounds such as high variability among components of one group, synergistic interactions between cysteine-knotted mini-proteins and other components (low molecular mass compounds and linear peptides), change of function from ion-channel acting mini-proteins to cytolytic effects and replacement of mini-proteins by linear peptides, acylpolyamines, large proteins or enzymes. We also add first phylogenetic considerations.

Brown recluse and black widow spiders.

"09/07"--Colophon.

The medical zoologist - Ronald Vernon Southcott

Dr Ronald Vernon Southcott (1918–1998) was amongst the greatest of the Australian doctor-naturalists. His toxinological contributions included the description and naming of the box-jellyfish, Chironex fleckeri, the first definitive study (1950–1957) of the toxinology, taxonomy and biology of Australian scorpions; and the first observations in Australia of the introduced fiddleback spider, Loxosceles. His research into the medical effects of toxic fungi, poisonous plants and Australian insects was extensive. He was a founding member of the International Society on Toxinology and served on the Toxicon Editorial Board for more than 30 years. He also made extensive contributions to acarology, and to the taxonomy of mites, specifically the sub-families and genera of the Erythraeoidea. This prodigious output was achieved by one who, with the exception of war service (1942–1946), almost never travelled outside South Australia, was almost entirely self-funded and worked from his home laboratory. With Dr. P.D. Scott and C.J. Glover, he was also the authority on the fish of South Australia. Dr. Southcott was also a medical epidemiologist and senior medical administrator (1949–1978) with the Australian Commonwealth Department of Veterans’ Affairs. He served for 30 years as an Honorary Consultant in Toxicology to the Adelaide Children's Hospital. As a zoologist and botanist of astounding breadth, he worked indefatigably in a voluntary capacity for the South Australian Museum, of which he was Museum Board Chairman from 1974 to 1982. In the pantheon of the great doctor-naturalists who have worked in Australia, he stands with Robert Brown and Thomas Lane Bancroft.