Computer-assisted Structural Analysis Of Regular Polysaccharides
Pure&App/. Chem., Vol. 61, No. 7, pp. 1181-1192 1989.Printed in Great Britain.@ 1989 IUPACComputer-assisted structural analysis of regularpolysaccharidesPer-Erik Jansson, Lennart Kenne and Goran WidmalmDepartment of Organic Chemistry, Arrhenius Laboratory, Universityof Stockholm, S-106 91 Stockholm, SwedenAbstract - A computer program, CASPER, for structural analysis ofoligo- and polysaccharides has been developed. Simple chemicalanalyses and NMR data (1H and/or 1 3 C NMR chemical shifts andl J C , H and 3 J H , H coupling constants) are used for determination ofthe structure.INTRODUCTIONCarbohydrates play an important role in Nature as e.g. energy source, buildingmaterial and in recognition processes. Due to the many different monosaccharidesexisting, the large number of combinations in which they can be linked, and theirability to form hydrophilic and hydrophobic regions, carbohydrates have manydifferent properties, and specific interactions with other molecules in biologicalsystems are common. To increase the understanding of their function in differentsurroundings and of the relationship between structure and biological activity, moreinformation on structural and physical properties of carbohydrates is of fundamentalimportance. To obtain relevant structural information on a carbohydrate molecule acomplete structural analysis with the following steps should be ofofofofofcomponentslinkagessequencepreferred conformationsthe dynamics of the moleculeIn Nature several different types of carbohydrates exist, e. g . bacterial polysaccharides, glycolipids, glycopeptides and different kinds of glycosides. Examples ofthese are given in Fig. 1. The bacterial polysaccharides are composed of oligosaccharide repeating units, in which a large number of different sugars could occur,many of them rare. Glycolipids and glycopeptides consist of a limited number ofdifferent sugars often occuring in similar oligosaccharide elements. These structuralelements can form larger oligosaccharides with a different numbers of branches andchain length. Saponins, an example among the glycosides, often contain two differentoligosaccharide chains linked to a triterpene or a sterol residue. In the structuralelucidation of these compounds both the structures of the two oligosaccharides andthe positions to which they are linked have to be determined.In the structural studies of carbohydrates a variety of methods have to be considered. The most used techniques today areSUGAR ANALYSISMETHYLATION ANALYSISCHEMICAL AND ENZYMIC DEGRADATIONSMASS SPECTROMETRYNMR SPECTROSCOPY1181
1182P.-E. JANSSON, L. KENNE AND G. WIDMALMa - L - Rh apc)1.13 3)-P-D-GlcpNAc-( 1 7)-P-Hepp-( l-t4t1P-D-GalpNAcHep Ip( 1-4)-P-D-GlcpNAc-( l - 2 ) a - D - M a n p ( 16)P-D-Manp( 1-4)-P-D-GlcpNAc-( l-.l)-P-D-GlcpNAc-( 1-N)-Asn3)u-NcuNAc-(2-6) -D-GaIp(l-4)- -D-GlcpNAc-(l-2)a-D-Manp(1’Fig. 1. Examples of different types of carbohydrates: a) 0-Polysaccharide fromVibrio cholerae 0:21; b) N -Acetyllactosamine type of carbohydrate chain fromantithrombin 111; c) Saponin from Nigella sativa.The sugar and methylation analyses are still of fundamental importance for thedetermination of components and linkages. The anomeric configurations can bedetermined with both enzymic and chemical reactions, optical rotation and NMRspectroscopy. Determination of the sequence has been a difficult and time-consumingtask and has mainly been performed by analysis of smaller oligosaccharides obtainedby enzymic or specific chemical degradations (ref. 1). Still today it is a difficultproblem and for each carbohydrate molecule different methods have to be useddepending upon structure and components of the polysaccharide. Mass spectrometryis now one of the most important methods for sequence analysis of oligosaccharides.The advantage with this technique is the high sensitivity which makes analysis ofsmall amounts of material possible but it is difficult to obtain information onanomeric configuration and linkage positions. Earlier only derivatized oligosaccharidescould be analysed, inserted into the spectrometer either via a gas chromatograph orvia a solid inlet interface. Recently soft-ionisation techniques, especially FAB-MS, hasbeen i n t r o d u c e d which e n a b l e s e q u e n c e d e t e r m i n a t i o n of underivatizedoligosaccharides. FAB-MS has been used in several structural investigations of oligoand polysaccharides (ref. 2).During the last years also NMR spectroscopy has become of increasing importance assuperconducting magnets, advanced computers, new multipulse experiments andtwo-dimensional techniques have become of general use. NMR spectroscopy is todayused routinely for analysis of components, substituents, linkages and sequences andthe structures of several oligo- and polysaccharides have been determined mainly orexclusively using one- or two-dimensional NMR techniques. Information from thechemical shift of structural reporter groups (ref. 3) and from nuclear Overhauserenhancements and scalar couplings over the glycosidic bond have been used toobtain sequence information. However, these techniques require expensive highfield NMR spectrometers and in most cases assignments of signals in the rathercomplex spectra, with several overlapping signals, are both difficult and timeconsuming.The efforts to develop new methods for future structural analysis of carbohydratesinvolve on one hand development of more sensitive techniques. These should enableanalysis of a large number of glycolipids and glycopeptides that are involved inimportant biological processes. On the other hand, development of faster and simplermethods are desirable as those mentioned are time-consuming and sometimes alsothe experiments are difficult to perform.
Computer-assisted structural analysis of regular polysaccharides1183A I M S FOR FUTURE STRUCTURAL ANALYSIS: LESS MATERIAL USEDSIMPLER ANALYSI SIn this paper a simpler and less time-consuming technique for sequence analysis ofoligo- and polysaccharides, using computer evaluation of NMR data and simplechemical analysis, will be presented. This technique is based on the assumption that1H and 13C NMR chemical shifts of the resonances from a monosaccharide residuewithin a larger saccharide depend mainly on the structure of the monosaccharideand on the nature of the flanking sugar residues. In the 1H NMR spectra most of thering protons have their resonances at similar chemical shifts forming a complexregion ( 6 3.5-4) with several overlapping signals. However, the resonances of someprotons appear outside this region. These protons have been named "Structuralreporter groups" including, inter alia, anomeric protons, deoxy protons and some ofthe equatorially disposed ring protons. As the chemical shifts of the correspondingsignals are dependent upon the components and linkages of the surrounding sugarresidues they could be correlated to the structure. Computerised approaches to thestructural analysis of oligo-saccharides from glycopeptides and glycolipids using the* H NMR chemical shifts of signals from the structural reporter groups have beenreported (refs. 4-6). The chemical shifts are compared with those for the signalsfrom corresponding residues in similar structures.A computerised approach to the structural analysis of unbranched regularpolysaccharides has been described (ref. 7). The method is based on evaluation ofthe 13C NMR spectra for all possible structures made up of the constituentmonosaccharides. It uses an additive scheme starting from the chemical shifts of the13C NMR resonances of the relevant monosaccharides to which the average values ofthe glycosylation shifts for signals from a- and P-carbons are added. This methodwas used in structural studies of some bacterial polysaccharides.U S E OF CASPER IN STRUCTURAL ANALYSIS OF OLIGO- A N DPOLYSACCHARIDESWe have described a computer program, CASPER, by which structural analysis oflinear polysaccharides with repeating units could be performed (ref. 8). Sugar andmethylation analysis data was used in combination with unassigned 13C NMRchemical shifts. The program can handle linear oligo- and polysaccharides composedof repeating units for which l3C NMR spectra for each possible permutation can besimulated and compared to the experimental spectrum. The simulation is based onan additivity approach, thus the chemical shift for each signal in the spectrum isassumed to be the sum of the chemical shift obtained from the monomeric residuesand a number of induced chemical shift changes, glycosylation shifts, which dependupon the linkage positions and the stereochemistry around the glycosidic bonds (Fig.2).'\OHFig. 2. The chemical shift values are thesum of the chemical shifts obtained fromthe monomer and the glycosylation shiftsfrom each flanking sugar.An improved and extended version of CASPER has recently been developed bywhich linear and branched structures can be analysed using one-dimensional 13CNMR data, two-dimensional H,H- or C,H-COSY data. In addition coupling constants foranomeric protons and carbons can be used. The program consists of a databaseandprocedures for generation of 1D and 2D spectra, for fitting spectra, forcomparison of J-values, for graphical display of spectra and an interface to themolecular modelling program CHEM-X.
P.-E. JANSSON, L. KENNE AND G. WIDMALM1184.**Welcome t EEEEEEEEERRRR*ASSSRw(R*AAAAAS PR RAAS PR R*CCCC AA SSSSPRR*Computer A s s i s t e d S p e c t r u m E v a l u a t i o n o f R e g u l a r p o l y s a c c h a r i d e s** Created b y G. W i d m a l m , P.-E. Jansson a n d L . Kenne. V e r s i o n 2 . 0 J u l y 1988.*******Top l e v e l menuI : I n f o r m a t i o n a b o u t CASPERD : Determination of structureR : Results of calculationG : Graphical outputS : Spectrum S i m u l a t i o n o f one s t r u c t u r eC : Chem-X 3D-fileE : End o f CASPER runEnter choice:Fig. 3. The banner and the choices inthe top level menu in CASPER.The database contains l H and l3C NMR chemical shifts for both anomeric forms ofeleven monosaccharides of which both aminosugars and uronic acids arerepresented. The glycosylation shifts for all 1H and 13C NMR resonances for all typesof disaccharide elements having pyranosidic rings, are included in the A6-file of thedatabase. These glycosylation shifts are mainly obtained from studies of 1,2-, 1,3-,1,4- and 1,6-linked disaccharides with different stereochemistry around theglycosidic bond (refs. 9-13).For some S h i g e l l a polysaccharides with branched 0-polysaccharides it was shownthat when the branched residue was not vicinally disubstituted additivity ofglycosylation shifts holds, but for vicinally disubstituted residues deviations arefound (ref. 14). To deal with such problems the extended version of CASPER containsa file with correction values for sugar residues in the branch point region. Thecorrection values are obtained from studies on "branched trisaccharides" (ref. 13).By the introduction of C,H-COSY spectra also the influence on the chemical shifts ofthe monosaccharide residue are considered. The basis for this is that the twodimensional information present in a pair of chemical shifts is lost on going to onedimension, i . e . , the one-dimensional spectrum. An example of this is the chemicalshifts for signals from H-3/C-3 and H-4/C-4 in P-Glc and in P-Gal, which pairwiseappear at 6 3.50176.6 and 3.42110.7 for P-Glc and 3.59173.8 and 3.89169.7 for P Gal. For the corresponding one-dimensional data the identification is less obvious.IIII'IFig. 4. The two-dimensional C/H-correlationcapsular polysaccharide from Klebsiella K 8 .spectrum of theSmith-degraded
Computer-assisted structural analysis of regular polysaccharides1185Another advantage with C,H-COSY spectra is the possibility to obtain all 1H NMRchemical shifts also for polysaccharides which normally give spectra of lowresolution and consequently less information. An example of this is given in Fig. 4.CIH-Correlation spectra has not been used extensively earlier because of the lowsensitivity of the 1 3 C nucleus. Recently, however, a new technique, "inversedetection" HMQC, utilizing detection of the 1H nucleus, has increased the sensitivityand made it possible to obtain C,H-COSY spectra of carbohydrates in considerablyshorter time (ref. 15).A flow diagram of the procedures for using CASPER in structural studies ofcarbohydrates is given below.Choice betweena)Structural determinationb)Simulation of one spectrumc)Generation of 3D-picturesIInput ofa)Components and linkagesChemical shifts; 6c or 6 or 6 / 6 b)c)J-valuesIGeneration of all possible structuresDeletion of J-incompatible structures (optional)ISimulation of spectraComparison of spectra, ranking of structuresIoutput ofa)Structuresb)Differences: experimental - simulatedc)Tabular display of sorted and non-sorted spectrad)Graphical display; 1-D spectra (*3C), 2-D-spectraTo use CASPER, data on components and linkage positions, normally obtained fromsugar and methylation analysis, are given to the program. All possible structures aregenerated by permutation of the components and addition of all combinations ofanomeric configurations. If the coupling constants for the anomeric protons, J H , H ,are available, these could be given to the program as the number of large (7-8 Hz),medium (3-4 Hz) and small ( 2 Hz) coupling constants refering to the normal valuesfor the j3-glucolgalacto, a - g l u c o l g a l a c t o and a l p - m a n n o configuration, respectively.Thus, the number 221 means that five sugar residues, two with p - g l u c o , two with a g l u c o and one with a - or p - m a n n o configuration, are the constituents of therepeating unit. The values for J c , Hof the signals from the anomeric carbons, ifavailable, are given as another number which refers to the size of the couplingconstant, ca 170 Hz - a , or ca 160 Hz - p. A number of 3 2 thus means three a - andtwo p -configurations. These J c , Hvalues also facilitate differentiation betweenresidues having a- and p - m a n n o configurations, as these have the same J H ,value,H 2 Hz. A comparison of the given numbers, representing the anomeric configurations,with the numbers from all suggested structures can then be performed. Thisprocedure drastically decreases the number of possible structures as all suggestedstructures with a non-corresponding set of anomeric configurations will be omitted.In the next step the program will simulate the 1H or 13C NMR spectra or the C,Hcorrelation spectra from all possible structures, using chemical shifts, glycosylationshifts and correction values from the database, and compare these with theexperimental spectrum. The fit, according to signal-to-signal comparison with theexperimental spectrum, is calculated and the simulated spectra are ranked accordingto their fit. The results from CASPER include inter alia a chosen number of suggestedstructures ranked according to their fit with the experimental spectrum. In additionto these, the As-sum, the deviationlsignal, the values for the coupling constants anda check number are given for each suggested structure. The AG-sum is the totalchemical shift difference between signals in the simulated spectrum and the
1186P.-E. JANSSON, L. KENNE AND G. WIDMALMexperimental spectrum. The structures are ranked according to this value. The checknumber describes the accuracy of the simulation, a low number is given when datafrom identical disaccharide elements or only minor approximations of these havebeen used. In addition the spectra can be shown and compared in the graphic modeas shown below.APPLICATIONS OF CASPER IN STRUCTURAL STUDIESThe use of CASPER in structural studies of carbohydrates will be examplified byanalysis of one oligosaccharide and three polysaccharides of known structure.In the first example a branched oligosaccharide will be analysed using unassigned13C NMR chemical shifts only. The oligosaccharide, which represents the repeatingunit in the 0-polysaccharide from Salmonella minnesota, consists of four sugarresidues of which a D -galactopyranosyl residue is branched. The oligosaccharide wassynthesized and analysed with 13C NMR spectroscopy by Kochetkov et al. (ref. 16)and the 13C NMR chemical shifts were corrected for differences in temperature andreference before giving them to CASPER (Fig. 5). In this example only the l3C NMRchemical shifts are used and the chemical shifts for signals from the a-form of thereducing residue were chosen. The results, obtained after permutation of thecomponents, generation of the anomeric configurations, simulation of spectra andcomparison of these with the experimental spectrum, are shown in Scheme 1, inwhich the four structures with the best fit are given. The correct structure has datawith the best fit but the structure in which the rhamnopyranosyl residue has a p configuration instead has only a somewhat larger AG-sum. If a comparison of thevalues for the coupling constants had been done structures 2 and 4 had beenomitted due to the incompatible IJC,H-values.The linear polysaccharide obtained by Smith-degradation of the capsularpolysaccharide from Klebsiella K8 consists of a trisaccharide repeating unit. Also inthis example only the unassigned l3C NMR chemical shifts were used in the analysis.As all residues are 3-linked only the differences in the anomeric configurations andin the stereochemistry around the linkages will be the factors influencing theglycosylation shifts. After permutation of the components and generation ofanomeric configurations the 13C NMR spectra of the suggested structures weresimulated.The results of the analysis is shown in Scheme 2. The A6-sum showing the totaldeviation is much smaller for the first suggested structure than for the otherstructures. In the simulated spectrum of the second structure (no 4) it is possible toobserve a large deviation for the signals from the anomeric carbons (Fig. 5). Thethird structure (no 6) has an incompatible number for the J-values and could also beomitted for that reason. The large A6-sums of the third and fourth structures showhow different the glycosylation shifts are when the substitution position is adjacentto an axial substituent as e . g . in 3-substituted galactopyranosyl residues.Experimental spectrum of SAL- MINFig. 5 . Graphical output of the experimental l3C NMR spectrum of the oligosacchariderepresenting the repeating unit in the 0-antigen from Salmonella minnesota and theanomeric region of the experimental and a simulated spectrum from the Smithdegraded capsular polysaccharide from Klebsiella K8.
Computer-assisted srrucrural analysis of regular polysaccharides1187SAL-MI NNo. Oligosaccharide.1BDMAN -4ALRHA -3ADGAL4ADGLC2BDMAN -4BLRHA -3ADGAL4ADGLC3BDMAN -4ADGALADGLC -4ALRHA4No.1234BDMAN -3ADGAL4ADGLC -4BLRHA13C13C H13CCheck#22222222312231220.210.210.210.3013C Experimental spectrum.102.3 101.3 100.8 93.2 79.173.4 73.1 72.6 71.7 71.362.1 61.5 61.3 18.078.1 77.371.3 70.3Spectrum number1.103.1 101.7 100.8 93.3 81.6 79.073.5 73.0 72.1 71.8 71.6 71.461.9 61.6 61.5 17.7Spectrum number101.7 101.2 100.373.4 73.1 72.961.9 61.4 61.4C-BranchCheck#0.010.010.010.0176.3 74.2 73.669.7 67.9 67.977.170.576.969.874.068.673.567.678.5 77.471.8 Scheme 1. Data from the four oligosaccharide structures with the best fit using 13CNMR data in CASPER. Calculations were performed for the oligosaccharide, whichrepresents the repeating unit of the 0-antigen of Salmonella minnesota.KL-K8SNo. GAL-3BDGAL13C13C JCH13CCheck#21021030021012123120.300.300.2113C Experimental spectrum.104.8 104.4 99.9 83.4 82.971.2 70.7 69.9 69.2 68.60.3080.0 76.461.8 61.775.6 73.161.571.4Spectrum number104.9 104.5 100.371.1 70.8 69.92.83.8 83.5 80.369.2 68.6 61.776.4 75.661.6 61.573.1 71.7Spectrum number105.0 104.1 95.971.2 70.5 71.6Scheme 2. Data from CASPER of the four structures for which the calculated 13C NMRspectra have the best fit with the experimental spectrum of the Smith-degradedcapsular poly-saccharide from Klebsiella K8.
P.-E. JANSSON, L. KENNE AND G. WIDMALM1188SH-FL-4 ANO.Polysaccharide.2 -3BDGLCN-2ALRHA -2ALRHA -3ALRHA6ADGLC-4 -3BDGLCN-2ALRHA -3ALRHA -2ALRHA-6ADGLC6 -3BDGLCN-3ALRHA -2ALRHA -2ALRHA 6ADGLC8 -6BDGLCN-2ALRHA -2ALRHA -3ALRHA3ADGLCNO.13C13C 220.322687.47.58.711.1 113 a n d JCH4512 -4 1 used t o eliminate structures174.9A103.01 0 1 . 8 h 1 0 1 . 8 101.675.2 74.1 73.3 73.2 72.870.7 70.0 69.9 69.9 69.417.5 .871.157.579.771.023.178.770.817.5S p e c t r u m number2.175.5 103.9 102.5 102.0 101.875.4 74.1 73.6 73.0 72.870.7 70.0 70.0 69.9 69.517.4 17.4S p e c t r u m number4.175.5 103.9 102.8 102.1 101.475.4 74.1 73.4 13.2 72.870.7 70.1 70.0 69.8 69.517.5 17.4S p e c t r u m 74.182.771.170.978.8S p e c t r u m 73.272.875.469.870.170.061.766.917.517.417.523.1 175.523.1 175.5Scheme 3. Data from CASPER of the four structures for which the calculated l3C NMR spectra havethe best fit with the experimental spectrum of Shigella flexneri type 4a 0-polysaccharide.For the branched structure of the repeating unit of Shigella flexneri type 4a, 1536possible structures were generated by permutation of the components and additionHare used to omit J of all combinations of the anomeric configuration. If J H ,valuesincompatible structures the number of possible strucmres decreases to 576. Afterthe same procedure with also the J c , H values only 288 possible structuresremained. Fitting the simulated spectra from these structures with the experimental13C NMR spectrum (Fig. 6) was then performed with the program.For the calculation of spectra no correction factors were used as there is no vicinaldisubstitution of the branching residue. In the program there are yet no specialglycosylation shifts for the acetamido group yet and this is partly the reason for therather high A6-sums obtained for all structures. Only minor differences in the A6sum are found between the four structures with the best fit. The only differences inthe structures are the sequence of the three a-L-rhamnopyranosyl residues (Scheme 3).
Computer-assisted structural analysis of regular polysaccharides1189To obtain the correct structure specific chemical reactions or NMR experiments haveto be performed.When using C,H-correlation data it is recommendable to use J-values for omittingstructures and simulation of the 13C NMR spectra must first be performed and onlythe structures with the best fit should be used in the fitting of C,H-correlationspectra as this procedure is time-consuming. The chemical shift difference is notgiven in ppm as a weighting factor is given to the 1H NMR chemical shifts for whichthere are smaller glycosylation shifts. An example of C,H-correlation simulations isdescribed for the Smith-degraded capsular polysaccharide from K l e b s i e l l a K8. Thesimulated spectra and a comparison with the experimental spectrum can also beshown in the graphic mode (Fig. 7).The structures with the best fit are the same as when using only 13C data but withdifferent order of the two last structures (Scheme 4). The AG-sums are higher thanin the 13C NMR simulation described above and are caused by the weighting factorused for the 1H NMR data.In the last example only 1H NMR data are used for calculation the structure of therepeating unit of the 0-polysaccharide from Shigella f l e x n e r i type Y. Thepolysaccharide is a linear tetrasaccharide repeat and the chemical shifts for thesignals have been determined using different H,H-COSY experiments (ref. 17). ForHtogetherthe calculations only information on com-ponents, linkages and J H ,valueswith the I H NMR chemical shifts for unassigned signals were given to the program.Structures with incompatible J H ,valuesHwere omitted before the calculations. The1H NMR chemical shifts for all possible structures were calculated and fitted to theexperimental data. The results could be displayed in the graphic mode (Fig. 8) or inthe tabular form (Scheme 5).IIIl 1lIIFig. 6. Experimental spectrum and corresponding graphical output in CASPER of theShigella flexneri type 4A polysaccharide.ImXi3.0R0'b4.0Xx0XFig. 7 . Graphical output of two-dimensional C/H-correlation spectra. Theexperimental spectrum of the Smith-degraded capsular polysaccharide fromK l e b s i e l l a K8 and the simulated spectra of the structures with best fit are shown.15.2
P.-E. JANSSON, L. KENNE AND G. WIDMALM1190KL-K8SNo. Polysaccharide.2 -3BDGLC -3BDGAL -3ADGAL4 -3BDGLC -3ADGAL -3BDGAL6 -3ADGLC -3BDGAL 300.300.210.3013.617.318.41H512 210 a n d JCH 12 u s e d t o e l i m i n a t e s t r u c t u r e sH , C - D e l t a s u m c a l c u l a t e d w i t h p o i n t of g r a v i t y a l g o r i t h mExperimental H,C-correlation spectrum.104.8 4.70104.4 4.7499.9 5.4080.0 4.0576.4 3 . 4 775.6 3.7271.2 3.8070.7 3.6869.9 4.2561.8 3.7661.8 3.7661.7 3.7661.5 .53.844.294.053.76S p e c t r u m number104.9 4.57104.580.3 3.9976.471.1 3.7770.861.7 3.7061.761.5 1.53.744.194.013.88S p e c t r u m number104.2 4.62104.278.7 3.1376.470.5 3.7269.861.7 3.7561.761.6 3.7185.873.968.161.73.743.594.013.7079.9 4.0471.4 4.1566.0 4.1261.6 3.90Scheme 4. Data from CASPER of the three structures for which the calculated C,Hcorrelation spectra have the best fit with the experimental spectrum.The first suggested structure is the repeating unit of the O-polysaccharide. In thenon-sorted data it is possible to see which of the simulated signals deviate from theexperimental values. The largest deviations are found for the signals from protonsclose to the N-acetyl group. In a molecular model of the two disaccharide elements(Fig. 9) containing the N-acetyl-P-D-glucosamineresidue interactions of the N-acetylgroup with neighbouring protons can be observed. Some correction factors for theinfluence of these interactions on the glycosylation shifts have been introduced.The l H NMR data can easily be obtained for the H-1 to H-4 signals of each residue ina polysaccharide, using different H,H-COSY experiments, and for many examples thisis enough to find the correct structure using CASPER. This makes it possible toanalyse polysaccharides of which there is only a small amount mu1aL.d .CII"I.9.2Fig. 8. Graphical output of the H,H-COSY spectrum from Shigella flexneri type Y polysaccharide. Each marker shows a certain spin-system (left) and spin-couplingconnectivities for ring protons are shown by different traces (right).1sPE13
1191Computer-assisted structural analysis of regular polysaccharidesSHIG-FL-Y-HNo. 0.881.06-3ALRHA-3BLRHA-2ALRHA-2BLRHA1 H #312231220.020.020.030.040.220.310.221.21512 1 0 3 u s e d t o e l i m i n a t e s t r u c t u r e s1H Experimental5.16 5.15 4.883.87 3.84 3.793.46 3.34 2.06spectrum.4.75 4.14 4.06 4.01 3.93 3.91 3.873.76 3.76 3.69 3.66 3.55 3.55 3.491.31 1 . 2 6 1 . 2 6S p e c t r u m number2.5.21 5.13 4.90 4.72 4 . 1 3 4.07 4.02 3.94 3.93 3.923.91 3.88 3.79 3.78 3.76 3.74 3.68 3.54 3.54 3.493.46 3.32 2.06 1.30 1.29 1.26S p e c t r u m number4.5 . 2 1 5.13 4 . 8 8 4.76 4.13 4 . 1 1 4.07 3.94 3 . 9 1 3.883.84 3.79 3.76 3.76 3.75 3.69 3.66 3.50 3.49 3.483 . 4 6 3 . 3 2 2 . 0 6 1.34 1 . 3 0 1 . 2 6S p e c t r u m number5.21 4.13 3.885.13 4.07 3.944.90 3.93 3.784.72 3.92 3.683.323.493.543.54S p e c t r u m number5.21 4.13 3.885.13 4.07 3.944.88 4 . 1 1 3.694.76 3.84 3.74 3 . 9 1 2.064.3.763.793.463.481.261.301.343.76 3.91 2.06Scheme 5. Data from CASPER on the four structures forwhich the calculated 1H NMR spectra have the best fitwith the experimental spectrum of Shigella flexneritype Y 0-polysaccharide.The non-sorted data forthe two structures with the best fit are also given.bFig. 9. Molecular models of disaccharideelements from the 0-antigen from S h i g cllri f l e x n e r i type Y.Inter-residueinteractions from the N-acetyl group aredepicted.PdFig. 10. Molecular model of the repeating unit of the Smith-degraded capsularpolysaccharide from K l e b s i e l l a K8. The model is produced by the molecularmodelling program CHEM-X, which is interfaced with CASPER, and the structuresobtained in CASPER can be built and studied in CHEM-X.
1192P.-E. JANSSON, L. KENNE AND G. WIDMALMIn CASPER there is also an interface to the molecular modelling program CHEM-X(ref. 18). The suggested structure will automatically be built using standardparameters and the structure could be displayed as a 3D-picture (Fig. 10).C O N C L U S I O N A N D FUTURE ASPECTSThe extension of CASPER to the use of more NMR parameters gives a largerpossibility to select the correct structure. For comp
USE OF CASPER IN STRUCTURAL ANALYSIS OF OLIGO- AND POLYSACCHARIDES We have described a computer program, CASPER, by which structural analysis of linear polysaccharides with repeating units could be performed (ref. 8). Sugar and methylation analysis data was used in combination with unassigned 13C NMR chemical shifts.
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VDSS Division of Family Services Assisted Living Facility Assessment Manual Assisted Living Facility (ALF) Any public or private assisted living facility that is required to be licensed as an assisted living facility by the Department of Social Services under Chapter 17 (§63.2-1700 et seq.) of Title .
2.1 Structural Health Monitoring Structural health monitoring is at the forefront of structural and materials research. Structural health monitoring systems enable inspectors and engineers to gather material data of structures and structural elements used for analysis. Ultrasonics can be applied to structural monitoring programs to obtain such .
12/16/2017 5136637 lopez damien 12/16/2017 5166979 lorenzano adam 12/16/2017 5117861 mejia martin 12/16/2017 5113853 milner gabriella 12/16/2017 5137867 navarro david 12/16/2017 5109380 negrete sylvia 12/16/2017 4793891 piliposyan alexander
