The long range goals of the proposed research is to understand how specificity signals encoded into protein sequences direct the co- and posttranstational processing of the linear polyamino acid products specified by the genetic code. It is becoming increasingly clear that co-/post-translational protein modification reactions represent key steps in the biosynthesis of biologically active proteins and in their transport to the appropriate compartments of action; our understanding of these reactions will be an important component in elucidating the genetic lesions in many inherited diseases and an absolute requirement for the full utilization of recombinant technology whenever proteins requiring extensive processing are produced in surrogate hosts lacking all or part of the appropriate processing apparatus. Two co-/post- translational reactions will be studied. The first, Glycan processing and recognition in glycoproteins, explores the fundamental question of how the protein matrix on which the common N-liked precursor glycan is located regulates the processing steps leading to different glycan products, and also how it affects the display of the glycan units in the interaction with receptors. Using neoglycoprotein models, the problem will be tackled by evaluating the substrate and ligand properties of well-defined free glycans in direct comparison with the same glycans associated with proteins in different types of complexes and with different glycan-protein stoichiometry. The second, the N-terminal processing of eukaryotic proteins, focuses on the reactions and the specificity signals involved in the well- established removal of the initiator methionine and the acylation of the new N-terminals amino acid in a large number of eukaryotic proteins. Building on the discovery of a muscle enzyme which catalyzes the specific removal of N- acetylmethionine from the N-terminus of short and long peptides, the proposed studies will focus on the enzymology of N-terminal trimming and acetylation in muscle and in yeast. The enzymes will be purified by conventional methods, and their specificity assessed by the use of different peptide (protein) substrates, the selection of which will be based on known acetylated and nonacetylated sequences in the data banks.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM031305-10
Application #
3279271
Study Section
Biochemistry Study Section (BIO)
Project Start
1982-03-01
Project End
1993-06-30
Budget Start
1991-07-01
Budget End
1993-06-30
Support Year
10
Fiscal Year
1991
Total Cost
Indirect Cost
Name
University of Texas Health Science Center Houston
Department
Type
Schools of Medicine
DUNS #
City
Houston
State
TX
Country
United States
Zip Code
77225
Shao, M C; Wold, F (1995) The effect of the protein matrix proximity on glycan reactivity in a glycoprotein model. Eur J Biochem 228:79-85
Krishna, R G; Chin, C C; Weldon, P J et al. (1995) Characterization of gamma-glutamyl transpeptidase from the Rathke's gland secretions of Kemp's ridley sea turtles (Lepidochelys kempi). Comp Biochem Physiol B Biochem Mol Biol 111:257-64
Sokolik, C W; Liang, T C; Wold, F (1994) Studies on the specificity of acetylaminoacyl-peptide hydrolase. Protein Sci 3:126-31
Shao, M C; Sokolik, C W; Wold, F (1994) Specificity studies of the GDP-[L]-fucose: 2-acetamido-2-deoxy-beta-[D]-glucoside (Fuc-->Asn-linked GlcNAc) 6-alpha-[L]-fucosyltransferase from rat-liver Golgi membranes. Carbohydr Res 251:163-73
Krishna, R G; Wold, F (1993) Post-translational modification of proteins. Adv Enzymol Relat Areas Mol Biol 67:265-98
Lu, Y; Ye, J; Wold, F (1993) Isolation of oligomannose-type glycans from bean glycoproteins. Anal Biochem 209:79-84
Chin, C C; Wold, F (1993) The use of tributylphosphine and 4-(aminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole in the study of protein sulfhydryls and disulfides. Anal Biochem 214:128-34
Krishna, R G; Wold, F (1992) Specificity determinants of acylaminoacyl-peptide hydrolase. Protein Sci 1:582-9
Shao, M C; Chin, C C (1992) Method for the detection of glycopeptides at the picomole level in HPLC peptide maps. Anal Biochem 207:100-5
Krishna, R G; Chin, C C; Wold, F (1991) N-terminal sequence analysis of N alpha-acetylated proteins after unblocking with N-acylaminoacyl-peptide hydrolase. Anal Biochem 199:45-50

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