Synthesis And Reactivity Of N-phosphono Amino Acids (NPAAs)

In this thesis,the reactivity of model compounds N-phosphono-amino acids(NPAAs) was explored for understanding the potential roles of P-N bond in bothprebiotic chemistry and biological chemistry.Cyclic acylphosphoramidates (2-hydroxy-2-oxy-1,3,2-oxazaphospholidine-5-ones,CAPAs) are phosphate activated amino acids possessing both a carboxyl-phosphorylanhydride and a phosphoramidate bond in a ring.The chemistry of CAPAs was almostunexplored since it was proposed in a prebiotic reaction of inorganic polyphosphates(PolyPs) with amino acids.In chapter 2,the bi-electrophilicity ofα-CAPAs(Gly-CAPA,Ala-CAPA) was identified by the isotope analysis (¹⁸O,¹⁵N) and furtherproved by trappingα-CAPA with nucleophiles such as water,amino acids,phosphateand methanol in alkaline media which yielded phosphorylated products,namely,theNPAAs and N-(mono-substituted)phosphoryl amino acids.To explore the property and reactivity of NPAAs needs fairly amount of pureNPAAs and their analogues.In chapter 3,first,directed by the CAPA involvedreaction mechanism between sodium trimetaphosphate (P₃m) and amino acids inchapter 2,a one-step reaction with efficient purification procedure in aqueous mediahas been developed for the synthesis of NPAAs.P₃m was used to phosphorylateamino acids to NPAAs with yields of 60～91%.The by-product inorganicpolyphosphates were recycled to regenerate the phosphorylation reagent P₃m.Then,for the preparation of N-phosphono and N-pyrophosphono derivatives of amineswithout an alpha carboxyl group,the method of P₃m/CaO was developed,in whichthe phosphate tail of N-triphosphoramidates was cleaved and precipitated by calciumion.Compounds from chapter 3 are studied by NMR-pH titration in chapter 4:pK_as ofphosphoryl oxygen and phosphoramidate nitrogen in a series of phosphoramidateswithout containing a vicinal carboxyl group were measured by ³¹p NMR-pH titration.However,the ³¹p NMR-pH titration curves ofα-NPAAs showed abnormal profile around pH range 0～5,which obscure the analysis of the site specific pKas of thecorresponding phosphate and carboxyl.The results revealed that pK_as betweenphosphate and carboxyl group inα-NPAAs are close to each other,which supportedthe proposed intramolecular H-bond between phosphate and carboxylate inα-NPAAs.In chapter 5,nucleophilic reactions of NPAAs in aqueous solution such ashydrolysis,N→N and N→O phosphate transfer reactions were investigated.It wasfound that neighboring functional groups such as carboxyl and hydroxyl groupcatalyzed the hydrolysis of NPAAs.In addition,the fragmentation pathways ofdifferent ionic species of NPAAs in gas phase was investigated by ESI tandem massspectrometry in both positive and negative modes.In chapter 6,the prebioticly available NPAAs were shown to be active in bothpeptides formation and phosphate transfer reaction.The processes involvesecond-order reactions in concentrated NPAAs solutions at near pH=7～8 at moderatetemperature.Phosphate transfer reaction between two molecular NPAAs leads toN-pyrophospho-amino acids,which spontaneously produce CAPAs for peptideformation.It was found that Mg²⁺ catalyzed the phosphate transfer from NPAAs tonucleoside monophosphates produce nucleoside diphosphates.It was suggested thatsuch non-enzymatic processes might have been existed on primitive earth.Based on the property and reactivity of NPAAs discovered in the thesis,a novelphosphate transfer mechanisms involving enzymatic N-phospholysine intermediate isproposed in chapter 7 which might provide some clues to the biological reactivitiesand roles of N-phospholysine.