3L05

Crystal structure of N-acetyl-L-ornithine transcarbamylase E92S mutant complexed with carbamyl phosphate and N-succinyl-L-norvaline

Replaces:  2G6A

Summary for 3L05

• Entry DOI: 10.2210/pdb3l05/pdb
• Related: 2FG7 3KZC 3KZK 3KZM 3KZN 3KZO 3L02 3L04 3L06
• Descriptor: N-acetylornithine carbamoyltransferase, N-(3-CARBOXYPROPANOYL)-L-NORVALINE, PHOSPHORIC ACID MONO(FORMAMIDE)ESTER, ... (5 entities in total)
• Functional Keywords: transcarbamylase, amino-acid biosynthesis, arginine biosynthesis, transferase
• Biological source: Xanthomonas campestris pv. campestris
• Cellular location: Cytoplasm : Q8P8J2
• Total number of polymer chains: 1
• Total formula weight: 40545.83

Authors

Shi, D.,Yu, X.,Allewell, N.M.,Tuchman, M. (deposition date: 2009-12-09, release date: 2010-03-31, Last modification date: 2023-11-22)

Primary citation

Shi, D.,Yu, X.,Cabrera-Luque, J.,Chen, T.Y.,Roth, L.,Morizono, H.,Allewell, N.M.,Tuchman, M.
A single mutation in the active site swaps the substrate specificity of N-acetyl-L-ornithine transcarbamylase and N-succinyl-L-ornithine transcarbamylase.
Protein Sci., 16:1689-1699, 2007

PubMed Abstract: Transcarbamylases catalyze the transfer of the carbamyl group from carbamyl phosphate (CP) to an amino group of a second substrate such as aspartate, ornithine, or putrescine. Previously, structural determination of a transcarbamylase from Xanthomonas campestris led to the discovery of a novel N-acetylornithine transcarbamylase (AOTCase) that catalyzes the carbamylation of N-acetylornithine. Recently, a novel N-succinylornithine transcarbamylase (SOTCase) from Bacteroides fragilis was identified. Structural comparisons of AOTCase from X. campestris and SOTCase from B. fragilis revealed that residue Glu92 (X. campestris numbering) plays a critical role in distinguishing AOTCase from SOTCase. Enzymatic assays of E92P, E92S, E92V, and E92A mutants of AOTCase demonstrate that each of these mutations converts the AOTCase to an SOTCase. Similarly, the P90E mutation in B. fragilis SOTCase (equivalent to E92 in X. campestris AOTCase) converts the SOTCase to AOTCase. Hence, a single amino acid substitution is sufficient to swap the substrate specificities of AOTCase and SOTCase. X-ray crystal structures of these mutants in complexes with CP and N-acetyl-L-norvaline (an analog of N-acetyl-L-ornithine) or N-succinyl-L-norvaline (an analog of N-succinyl-L-ornithine) substantiate this conversion. In addition to Glu92 (X. campestris numbering), other residues such as Asn185 and Lys30 in AOTCase, which are involved in binding substrates through bridging water molecules, help to define the substrate specificity of AOTCase. These results provide the correct annotation (AOTCase or SOTCase) for a set of the transcarbamylase-like proteins that have been erroneously annotated as ornithine transcarbamylase (OTCase, EC 2.1.3.3).

PubMed: 17600144
DOI: 10.1110/ps.072919907

Experimental method

X-RAY DIFFRACTION (2.8 Å)