Importance of non-conserved distal carboxyl terminal amino acids in two peptidases belonging to the M1 family: Thermoplasma acidophilum Tricorn interacting factor F2 and Escherichia coli Peptidase N

Kumar, Anujith and Bhosale, Manoj and Reddy, Surendranath and Srinivasan, Narayanaswamy and Nandi, Dipankar (2009) Importance of non-conserved distal carboxyl terminal amino acids in two peptidases belonging to the M1 family: Thermoplasma acidophilum Tricorn interacting factor F2 and Escherichia coli Peptidase N. In: Biochimie, 91 (9). pp. 1145-1155.

PDF http___www.sciencedirect.com_science__ob=MImg&_imagekey=B6VRJ-4WHFD9J-2-3&_cdi=6236&_user=512776&_orig=search&_coverDate=09%2F30%2F2009&_sk=999089990&view=c&wchp=dGLbVlb-zSkWA&md5=2a66f9bef89fa.pdf - Published Version Restricted to Registered users only Download (3MB) | Request a copy

Abstract

Enzymes belonging to the M1 family play important cellular roles and the key amino acids (aa) in the catalytic domain are conserved. However, C-terminal domain aa are highly variable and demonstrate distinct differences in organization. To address a functional role for the C-terminal domain, progressive deletions were generated in Tricorn interacting factor F2 from Thermoplasma acidophilum (F2) and Peptidase N from Escherichia coli (PepN). Catalytic activity was partially reduced in PepN lacking 4 C-terminal residues (PepNΔC4) whereas it was greatly reduced in F2 lacking 10 C-terminal residues (F2ΔC10) or PepN lacking eleven C-terminal residues (PepNΔC11). Notably, expression of PepNΔC4, but not PepNΔC11, in E. coliΔpepN increased its ability to resist nutritional and high temperature stress, demonstrating physiological significance. Purified C-terminal deleted proteins demonstrated greater sensitivity to trypsin and bound stronger to 8-amino 1-napthalene sulphonic acid (ANS), revealing greater numbers of surface exposed hydrophobic aa. Also, F2 or PepN containing large aa deletions in the C-termini, but not smaller deletions, were present in high amounts in the insoluble fraction of cell extracts probably due to reduced protein solubility. Modeling studies, using the crystal structure of E. coli PepN, demonstrated increase in hydrophobic surface area and change in accessibility of several aa from buried to exposed upon deletion of C-terminal aa. Together, these studies revealed that non-conserved distal C-terminal aa repress the surface exposure of apolar aa, enhance protein solubility, and catalytic activity in two soluble and distinct members of the M1 family.

Item Type:	Journal Article
Publication:	Biochimie
Publisher:	Elsevier Science
Additional Information:	Copyright of this article belongs to Elsevier Science.
Keywords:	C-Terminal domain;Cytosolic protein degradation;Enzyme solubility and function;Metallopeptidase;M1 family.
Department/Centre:	Division of Biological Sciences > Molecular Biophysics Unit Division of Biological Sciences > Biochemistry
Date Deposited:	16 Dec 2009 07:05
Last Modified:	19 Sep 2010 05:46
URI:	http://eprints.iisc.ac.in/id/eprint/23486

Actions (login required)

View Item