50 years of Mass Spectrometry at Rudjer Boskovic Institute

Dan je pregled aktivnosti i znanstvenih rezultata suradnika Laboratorija za kemijsku kinetiku (kasnije Laboratorija za kemijsku kinetiku i atmosfersku kemiju) Instituta Ruđer Bošković na području spektrometrije masa od 1959. do danas.An overview is given of activities and scientific results by coworkers of the Laboratory for Chemical Kinetics (later Laboratory for Chemical Kinetics and Atmospheric Chemistry) of the Rudjer Boskovic Institute in the field of mass spectrometry from 1959 to the present

50 years of Mass Spectrometry at Rudjer Boskovic Institute

Dan je pregled aktivnosti i znanstvenih rezultata suradnika Laboratorija za kemijsku kinetiku (kasnije Laboratorija za kemijsku kinetiku i atmosfersku kemiju) Instituta Ruđer Bošković na području spektrometrije masa od 1959. do danas.An overview is given of activities and scientific results by coworkers of the Laboratory for Chemical Kinetics (later Laboratory for Chemical Kinetics and Atmospheric Chemistry) of the Rudjer Boskovic Institute in the field of mass spectrometry from 1959 to the present

High-throughput discrimination of bacteria isolated from Astacus astacus and A. leptodactylus

Bacterial diseases and pathogens of crayfish are common, widespread, and occasionally causing serious mortalities. In order to take rapid measures for correct treatment of crayfish diseases, the turnover time and accuracy in bacterial identification is an issue. Bacteria isolated from tissues of apparently healthy Astacus astacus and A. leptodactylus were identified by the commercial phenotypic tests (API 20E) and by the matrix assisted laser induced desorption ionization connected to the time of flight mass spectrometry (MALDI-TOF MS). For Gram-negative rods, API 20E resulted in fewer species identifications than MALDI-TOF MS (5.2% versus 52.61%). The most frequently identified genus from A. astacus and A. leptodactylus was Pseudomonas spp.: API 20E (47.82%) and MALDI-TOF MS (52.17%). Both systems identified 60.86% of total isolates identically to the genus. Hafnia alvei was the only isolate for which API 20E and MALDI-TOF MS had a concordant reading to the species. MALDI-TOF MS proved to be a powerful, low-cost, rapid tool in bacterial genus identification. This is the first report of a direct comparison between the two systems for the identification of bacteria in crayfish, and also the first report on using MALDI-TOF MS for discrimination of freshwater crayfish bacterial isolates

Gas-phase metallation reactions of porphyrins with metal monocations

The gas-phase reactions of octaethylporphyrin (H 2P, III) with 12 metal monocations, which were produced by laser ablation of Al, V, Cr, Fe, Ni, Nb, Mo, Hf, W, Re, Pt, and Au targets in an ICR-FTMS instrument, revealed three possible reaction pathways: (i) charge transfer; (ii) metal incorporation (ligation) with loss of H 2; and (iii) dimerization of the ligation product from pathway ii by addition of another porphyrin without further loss of H 2. The differences in metal behavior are discussed. The reaction of a mixture of hexaethyldimethylporphyrin (H 2P, I) and pentaethyltrimethylporphyrin (H 2P, II) with neat Fe + was also investigated; it was found that more than half of the Fe + reacts by charge exchange and that the lower-alkylated porphyrin (I), which has the higher ionization energy (IE), reacts more slowly in charge exchange but with the same rate in the both ligation and dimerization reactions as the higher alkylated porphyrin (H 2P, II)

Particulate sulfur-containing lipids: Production and cycling from the epipelagic to the abyssopelagic zone

There are major gaps in our understanding of the distribution and role of lipids in the open ocean especially with regard to sulfur-containing lipids (S-lipids). Here, we employ a powerful analytical approach based on high resolution Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) to elucidate depth-related S-lipid production and molecular transformations in suspended particulate matter from the Northeast Atlantic Ocean in this depth range. We show that within the open-ocean environment S-lipids contribute up to 4.2% of the particulate organic carbon, and that up to 95% of these compounds have elemental compositions that do not match those found in the Nature Lipidomics Gateway database (termed “novel”). Among the remaining 5% of lipids that match the database, we find that sulphoquinovosyldiacylglycerol (SQDG) are efficiently removed while sinking through the mesopelagic zone. The relative abundance of other assigned lipids (sulphoquinovosylmonoacylglycerol (SQMG), sulfite and sulfate lipids, Vitamin D2 and D3 derivatives, and sphingolipids) did not change substantially with depth. The novel S-lipids, represented by hundreds of distinct elemental compositions (160–300 molecules at any one depth), contribute increasingly to the lipid and particulate organic matter pools with increased depth. Depth-related transformations cause (i) incomplete degradation/transformation of unsaturated S-lipids which leads to the depth-related accumulation of the refractory saturated compounds with reduced molecular weight (average 455 Da) and (ii) formation of highly unsaturated S-lipids (average abyssopelagic molecular double bond equivalents, DBE=7.8) with lower molecular weight (average 567 Da) than surface S-lipids (average 592 Da). A depth-related increase in molecular oxygen content is observed for all novel S-lipids and indicates that oxidation has a significant role in their transformation while (bio)hydrogenation possibly impacts the formation of saturated compounds. The instrumentation approach applied here represents a step change in our comprehension of marine S-lipid diversity and the potential role of these compounds in the oceanic carbon cycle. We describe a very much higher number of compounds than previously reported, albeit at the level of elemental composition and fold-change quantitation with depth, rather than isomeric confirmation and absolute quantitation of individual lipids. We emphasize that saturated S-lipids have the potential to transfer carbon from the upper ocean to depth and hence are significant vectors for carbon sequestration

A novel Porphyromonas gingivalis enzyme: An atypical dipeptidyl peptidase III with an ARM repeat domain.

Porphyromonas gingivalis, an asaccharolytic Gram-negative oral anaerobe, is a major pathogen associated with adult periodontitis, a chronic infective disease that a significant percentage of the human population suffers from. It preferentially utilizes dipeptides as its carbon source, suggesting the importance of dipeptidyl peptidase (DPP) types of enzyme for its growth. Until now DPP IV, DPP5, 7 and 11 have been extensively investigated. Here, we report the characterization of DPP III using molecular biology, biochemical, biophysical and computational chemistry methods. In addition to the expected evolutionarily conserved regions of all DPP III family members, PgDPP III possesses a C-terminal extension containing an Armadillo (ARM) type fold similar to the AlkD family of bacterial DNA glycosylases, implicating it in alkylation repair functions. However, complementation assays in a DNA repair-deficient Escherichia coli strain indicated the absence of alkylation repair function for PgDPP III. Biochemical analyses of recombinant PgDPP III revealed activity similar to that of DPP III from Bacteroides thetaiotaomicron, and in the range between activities of human and yeast counterparts. However, the catalytic efficiency of the separately expressed DPP III domain is ~1000-fold weaker. The structure and dynamics of the ligand-free enzyme and its complex with two different diarginyl arylamide substrates was investigated using small angle X-ray scattering, homology modeling, MD simulations and hydrogen/deuterium exchange (HDX). The correlation between the experimental HDX and MD data improved with simulation time, suggesting that the DPP III domain adopts a semi-closed or closed form in solution, similar to that reported for human DPP III. The obtained results reveal an atypical DPP III with increased structural complexity: its superhelical C-terminal domain contributes to peptidase activity and influences DPP III interdomain dynamics. Overall, this research reveals multifunctionality of PgDPP III and opens direction for future research of DPP III family proteins.</p

High-throughput discrimination of bacteria isolated from Astacus astacus and A. leptodactylus

Bacterial diseases and pathogens of crayfish are common, widespread, and occasionally causing serious mortalities. In order to take rapid measures for correct treatment of crayfish diseases, the turnover time and accuracy in bacterial identification is an issue. Bacteria isolated from tissues of apparently healthy Astacus astacus and A. leptodactylus were identified by the commercial phenotypic tests (API 20E) and by the matrix assisted laser induced desorption ionization connected to the time of flight mass spectrometry (MALDI-TOF MS). For Gram-negative rods, API 20E resulted in fewer species identifications than MALDI-TOF MS (5.2% versus 52.61%). The most frequently identified genus from A. astacus and A. leptodactylus was Pseudomonas spp.: API 20E (47.82%) and MALDI-TOF MS (52.17%). Both systems identified 60.86% of total isolates identically to the genus. Hafnia alvei was the only isolate for which API 20E and MALDI-TOF MS had a concordant reading to the species. MALDI-TOF MS proved to be a powerful, low-cost, rapid tool in bacterial genus identification. This is the first report of a direct comparison between the two systems for the identification of bacteria in crayfish, and also the first report on using MALDI-TOF MS for discrimination of freshwater crayfish bacterial isolates