| Description |
This dissertation is composed of three major topics: the structure determination, principally by mass spectrometry, of a new nucleotide antibiotic, phosmidosine; a comparative study of high and low energy collision-induced dissociation (CID) of dinucleoside monophosphate ions; and the results of which are applied to the identification of 2'-O-methyllribonucleotidylyl ribonucleosides formed by ribonuclease T2 digestion of tRNA. The structure determination of phosmidosine was complicated by chemical instability and the presence of impurities in the isolate. The necessity of isolating the phosmidosine was circumvented by using tandem mass spectrometry (MS/MS), and liquid chromatography-mass spectrometry (LC/MS). The majority of MS/MS experiments were performed on the isopropylidene derivative because of increase sample ion abundance from fast atom bombardment (FAB). The structure of the nucleoside, produced from phosimdosine by base hydrolysis, was confirmed by comparing LC/MS and gas chromatography-mass spectrometry data to that of synthetic 8-hydroxyadenosine. The phosmidosine isolate also contained two homologs and an isomer that were characterized. Although there have been studies of high energy CID of negative ions from dinucleoside monophosphates, previous examination of positive or negative ions from dinucleoside monophosphates by low energy CID has not been reported. Low energy CID spectra were obtained for all common heterodinucleoside monophosphates and three 2'-O-methlribonucleotidylyl ribonucleosides. It was established that a dinucleoside monophosphate sequence could be determined by comparing nucleoside base ion peak height in the CID mass spectra of (M-H)¯, and that ion m/z 81 (C5H5O+) from deoxyribo- or m/z 97 (C5H5O+) from ribodinucleoside monophosphates, predominantly produced from the 3'-terminus nucleoside sugar, is observed in all CID mass spectra of MH+ from dinucleoside monophosphates. 2'-O-Methylribonucleotidylyl found in RNase T2 digested RNA can be identified directly from the digest by using precursor ion scans of m/z 97, and product ion mass spectra provide conformation and sequence information. This is of interest because 2'-O-methylribonucleotidylyl ribonucleosides often poses an special problem for RNA sequencing because of their stability against chemical and enxymatic hydrolysis. |
