Jeffrey L Urbauer
The University of Georgia, USA
Title: Oxidative stress, methionine oxidation, and calmodulin structure and function
Biography
Biography: Jeffrey L Urbauer
Abstract
Statement of the Problem: Oxidation of methionine residues in proteins to methionine sulfoxide is a prevalent, reversible post-translational modification. Changes in protein structure and function accompany oxidation due to polarity and steric differences between methionine and the sulfoxide. We are investigating the consequences of methionine oxidation in the regulatory protein calmodulin (CaM), a key calcium signal transducer with nine methionine residues, in hydrophobic pockets of its opposing globular domains, which interact with target proteins. CaM with oxidized methionine residues accumulates under conditions of oxidative stress, and because of its central role in biology, it is important to understand the functional effects of these alterations and their physical origins. Methodology: Methionine residues in CaM are easily oxidized in vitro with hydrogen peroxide. To study the effects of oxidation of specific methionine residues, leucine was substituted for methionine at remaining sites. A combination of functional assays, single molecule studies, and NMR spectroscopy were used to assess functional and structural consequences of methionine oxidation. Findings: For the best studied case, activation of the plasma membrane Ca-ATPase (PMCA) by CaM, impaired CaM function is due to oxidation of a single C-terminal methionine. Single molecule experiments indicate non-productive binding of oxidized CaM to the PMCA. High resolution NMR studies demonstrate significant structural perturbation in the C-terminal globular domain of oxidized CaM and an inability to anchor the PMCA to this domain. Conclusion & Significance: The functional effects of methionine oxidation in CaM are highly target dependent, as is the degree to which selective oxidation of particular methionine residues in CaM affects function. The results of CaM activation of the PMCA also indicate that both high-affinity productive and non-productive complexes of oxidized CaM with targets are possible. These facts indicate that a comprehensive understanding of the metabolic consequences of CaM oxidation will be challenging.
References:
- Bartlett RK, Bieber Urbauer RJ, Anbandam A, Smallwood HS, Urbauer JL, Squier TC (2003) Oxidation of Met144 and Met145 in calmodulin blocks calmodulin dependent activation of the plasma membrane ca-ATPase. Biochemistry 42:3231-3238.
- Osborn KD, Bartlett RK, Mandal A, Zaidi A, Bieber Urbauer RJ, Urbauer JL, Galeva N, Williams TD, Johnson CK (2004) Single-molecule dynamics reveal an altered conformation for the autoinhibitory domain of plasma membrane Ca-ATPase bound to oxidatively modified calmodulin. Biochemistry 43:12937-12944.
- Anbanandam A, Bieber Urbauer RJ, Bartlett RK, Smallwood HS, Squier TC, Urbauer, JL (2005) Mediating molecular recognition by methionine oxidation: Conformational switching by oxidation of methionine in the carboxyl-terminal domain of calmodulin. Biochemistry 44:9486-9496.
- Slaughter BD, Bieber Urbauer RJ, Urbauer JL, Johnson CK (2007) Mechanism of calmodulin recognition of the binding domain of isoform 1b of the plasma membrane Ca-ATPase: Kinetic pathway and effects of methionine oxidation. Biochemistry 46: 4045-4054.
- Lubker C, Bieber Urbauer RJ, Moskovitz J, Dove S, Weisemann J, Fedorova M, Urbauer JL, Seifert R (2015) Membranous adenylyl cyclase 1 activation is regulated by oxidation of N- and C-terminal methionine residues in calmodulin. Biochemical Pharmacology 93:196-209.