Biochemical and Structural Studies of Pseudouridine 55 Synthase

Sequence alignment of the families of PsiS identifies the conserved aspartic acid discussed previously as the strictly conserved amino acid. Structural alignment identifies the conserved tyrosine, which is part of hydrophobic core in the active site. The role of the conserved tyrosine is less obvious in addition to apparent structural role. Enzymatic activity assay reveals that mutating T. maritima Tyr67 to any other amino acids abolishes the enzymatic activity. Structure of T. maritima Y67F in complex with the 5FU-RNA reveals, however, that the same 5FhPsi product formed by wild-type Psi55S is also found in the active site. Furthermore, HPLC analysis indicates that 5FhPsi is also formed when 5FU RNA was incubated with either E. coli Y76F or Y76L but not with Y76A. The combined information from structural, biochemical and mutational studies allows us to propose the likely role of the conserved tyrosine as a general base for proton abstraction in PsiS-catalyzed reaction as well as structural role of the hydrophobic phenol ring in the active site.

Name thesis:degree_name

Ph.D.

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biochemistry

Grantor

University of Illinois at Urbana-Champaign

Year dc:date

2015

Author dc:creator

• Phannachet, Kulwadee

Contributors dc:contributor

• Huang, Raven H.

• Chemistry, Biochemistry

Language dc:language

eng

Identifier

(MiAaPQ)AAI3182354

OAI identifier oai:identifier

oai:www.ideals.illinois.edu:2142/84812