This invention provides compositions and methods for producing translational components that expand the number of genetically encoded amino acids in eukaryotic cells. The components include orthogonal tRNAs, orthogonal aminoacyl-tRNA synthetases, orthogonal pairs of tRNAs/synthetases and unnatural amino acids. Proteins and methods of producing proteins with unnatural amino acids in eukaryotic cells are also provided.

The present invention provides a process of producing a trifluoromethoxylated aryl or trifluoromethoxylated heteroaryl having the structure:

##STR00001##

wherein A is an aryl or heteroaryl, each with or without subsutitution; and R.sub.1 is —H, -(alkyl), -(alkenyl), -(alkynyl), -(aryl), -(heteroaryl), -(alkylaryl), -(alkylheteroaryl), —NH-(alkyl), —N(alkyl).sub.7, —NH-(alkenyl), —NH-(alkynyl) —NH-(aryl), —NH-(heteroaryl), —O-(alkyl), —O-(alkenyl), —O-(alkynyl), —O-(aryl), —O-(heteroaryl), —S-(alkyl), —S-(alkenyl), —S-(alkynyl), —S-(aryl), or —S-(heteroaryl), comprising: (a) reacting a compound having the structure:

##STR00002##

with a trifluoromethylating agent in the presence of a base in a first suitable solvent under conditions to produce a compound having the structure:

##STR00003##

and (b) maintaining the compound produced in step (a) in a second suitable solvent under conditions sufficient to produce the trifluoromethoxylated aryl or trifluormethoxylated heteroaryl having the structure:

##STR00004##

The present invention provides a process of producing a trifluoromethoxylated aryl or trifluoromethoxylated heteroaryl having the structure:

##STR00001##

wherein A is an aryl or heteroaryl, each with or without subsutitution; and R.sub.1 is —H, -(alkyl), -(alkenyl), -(alkynyl), -(aryl), -(heteroaryl), -(alkylaryl), -(alkylheteroaryl), —NH-(alkyl), —N(alkyl).sub.7, —NH-(alkenyl), —NH-(alkynyl) —NH-(aryl), —NH-(heteroaryl), —O-(alkyl), —O-(alkenyl), —O-(alkynyl), —O-(aryl), —O-(heteroaryl), —S-(alkyl), —S-(alkenyl), —S-(alkynyl), —S-(aryl), or —S-(heteroaryl), comprising: (a) reacting a compound having the structure:

##STR00002##

with a trifluoromethylating agent in the presence of a base in a first suitable solvent under conditions to produce a compound having the structure:

##STR00003##

and (b) maintaining the compound produced in step (a) in a second suitable solvent under conditions sufficient to produce the trifluoromethoxylated aryl or trifluormethoxylated heteroaryl having the structure:

##STR00004##

A method for at least partial deacetylation of a biopolymer comprising acetyl groups, including: a1) providing a biopolymer including acetyl groups; a2) reacting the biopolymer including acetyl groups with hydroxylamine (NH.sub.2OH) or a salt thereof at a temperature of 100° C. or less for 2-200 hours to form an at least partially deacetylated biopolymer; and a3) recovering the at least partially deacetylated biopolymer.

A method for at least partial deacetylation of a biopolymer comprising acetyl groups, including: a1) providing a biopolymer including acetyl groups; a2) reacting the biopolymer including acetyl groups with hydroxylamine (NH.sub.2OH) or a salt thereof at a temperature of 100° C. or less for 2-200 hours to form an at least partially deacetylated biopolymer; and a3) recovering the at least partially deacetylated biopolymer.

The invention generally relates to an improved process for the preparation of N-((1R,2S,5R)-5-(tert-butylamino)-2-((S)-3-(7-tert-butylpyrazolo[1,5-a][1,3,5]triazin-4-ylamino)-2-oxopyrrolidin-1-yl)cyclohexyl)acetamide, ##STR00001## as well as novel intermediates employed in the process, which may be useful for the treatment of cancer.

The invention generally relates to an improved process for the preparation of N-((1R,2S,5R)-5-(tert-butylamino)-2-((S)-3-(7-tert-butylpyrazolo[1,5-a][1,3,5]triazin-4-ylamino)-2-oxopyrrolidin-1-yl)cyclohexyl)acetamide, ##STR00001## as well as novel intermediates employed in the process, which may be useful for the treatment of cancer.

Methods and reagents for esterification of biological molecules including proteins, polypeptides and peptides. Diazo compounds of formula I: ##STR00001##
where R is hydrogen, an alkyl, an alkenyl or an alkynyl, R.sub.A represents 1-5 substituents on the indicated phenyl ring and R.sub.M is an organic group, which includes a label, a cell penetrating group, a cell targeting group, or a reactive group or latent reactive group for reaction to bond to a label, a cell penetrating group, or a cell targeting group, among other organic groups are useful for esterification of biological molecules. Also provided are diazo compounds which are bifunctional and trifunctional coupling reagents as well as reagents for the synthesis of compounds of formula I.

Methods and reagents for esterification of biological molecules including proteins, polypeptides and peptides. Diazo compounds of formula I: ##STR00001##
where R is hydrogen, an alkyl, an alkenyl or an alkynyl, R.sub.A represents 1-5 substituents on the indicated phenyl ring and R.sub.M is an organic group, which includes a label, a cell penetrating group, a cell targeting group, or a reactive group or latent reactive group for reaction to bond to a label, a cell penetrating group, or a cell targeting group, among other organic groups are useful for esterification of biological molecules. Also provided are diazo compounds which are bifunctional and trifunctional coupling reagents as well as reagents for the synthesis of compounds of formula I.

Methods and reagents for esterification of biological molecules including proteins, polypeptides and peptides. Diazo compounds of formula I: ##STR00001##
where R is hydrogen, an alkyl, an alkenyl or an alkynyl, R.sub.A represents 1-5 substituents on the indicated phenyl ring and R.sub.M is an organic group, which includes a label, a cell penetrating group, a cell targeting group, or a reactive group or latent reactive group for reaction to bond to a label, a cell penetrating group, or a cell targeting group, among other organic groups are useful for esterification of biological molecules. Also provided are diazo compounds which are bifunctional and trifunctional coupling reagents as well as reagents for the synthesis of compounds of formula I.