Slippery Substrates Impair ATP-dependent Protease Function by Slowing Unfolding

THE JOURNAL OF BIOLOGICAL CHEMISTRY VOL. 289, NO. 6, p. 3826, February 7, 2014 Published in the U.S.A. LETTER Slippery Substrates Impair ATP-dependent Protease Function by Slowing Unfolding The first 7 figures in the article by Too et al. (1) consider the compo- sition of substrates that cause an ATPase motor to slip and the topo- logic escape mode of intermediates. This lab previously showed that amino acid sequences (of viral origin) containing only glycine and alanine residues can cause stalling of proteasome degradation and processivity failure. Production of degradation intermediate end products is promoted by the joint action of a hard-to-unfold domain and a sequence that impairs delivering the translocation force that impels unfolding (3, 4). In Ref. 1 we asked whether the peculiar sequence elaborated by a viral human pathogen can also thwart a 3826 JOURNAL OF BIOLOGICAL CHEMISTRY Philip Coffino 1 , Priscilla Hiu-Mei Too 2 , and Jenny Erales 3 Department of Microbiology and Immunology, University of Califor- nia, San Francisco, California 94143 1. Too, P. H., Erales, J., Simen, J. D., Marjanovic, A., and Coffino, P. (2013) Slippery substrates impair function of a bacterial protease ATPase by unbalancing translocation versus exit. J. Biol. Chem. 2. Kraut, D. A. (2013) Slippery substrates impair ATP-dependent protease function by slowing unfolding. J. Biol. Chem. 288, 34729 –34735 3. Hoyt, M. A., Zich, J., Takeuchi, J., Zhang, M., Govaerts, C., and Coffino, P. (2006) Glycine-alanine repeats impair proper substrate unfolding by the proteasome. EMBO J. 25, 1720 –1729 4. Zhang, M., and Coffino, P. (2004) Repeat sequence of Epstein-Barr virus-encoded nuclear antigen 1 protein interrupts proteasome substrate processing. J. Biol. Chem. 279, 8635– 8641 5. Kraut, D. A., Israeli, E., Schrader, E. K., Patil, A., Nakai, K., Nanavati, D., Inobe, T., and Matouschek, A. (2012) Sequence- and species-dependence of proteasomal processivity. ACS Chem. Biol. 7, DOI 10.1074/jbc.L113.532622 E-mail: philip.coffino@ucsf.edu Present address: 688 Minor Hall, University of California, Berkeley, CA 94702. Present address: Campus de Luminy, 163 Ave. de Luminy, 13288 Marseille CEDEX 09, France. VOLUME 289 • NUMBER 6 • FEBRUARY 7, 2014 Downloaded from http://www.jbc.org/ at UCSF Library & CKM on March 25, 2015 Analysis of data in Fig. 8 of Too et al. (1) was flawed by our naive error, now corrected by Kraut (2). In fitting the first order rate constant k proc for processive degradation by the complex between ClpXP and a stalled substrate, we ignored simultaneous decay by a second process, exit of substrate. Modeling by Kraut (2) now supplies accurate values of pertinent rate constants and reverses a conclusion of Too et al. (1). Slippery sequences predominantly change processive unfolding kinetics, not substrate dissociation. We here observe science advanc- ing as it should: attentively interactive, self-correcting. bacterial ATPase. It can. Systematic investigation of the composi- tion of sequences that cause slippage revealed that side chains of simple shape and small size promote polypeptide slipping. This result contrasts markedly with an alternate view, e.g. in Ref. 5, that regions of “low sequences complexity” produce slipping. Persistent use of “sequence complexity” terminology in Ref. 2 to describe slippery sequences dismisses and fails to engage the major findings of Too et al.