Amino acid racemization reveals differential protein turnover in osteoarthritic articular and meniscal cartilages

Thomas V Stabler,Robert D Zura,Virginia Kraus,Samuel S Byers

doi:10.1186/ar2639

Abstract

IntroductionCertain amino acids within proteins have been reported to change from the L form to the D form over time. This process is known as racemization and is most likely to occur in long-lived low-turnover tissues such as normal cartilage. We hypothesized that diseased tissue, as found in an osteoarthritic (OA) joint, would have increased turnover reflected by a decrease in the racemized amino acid content.MethodsUsing high-performance liquid chromatography methods, we quantified the L and D forms of amino acids reported to racemize in vivo on a biological timescale: alanine, aspartate (Asp), asparagine (Asn), glutamate, glutamine, isoleucine, leucine (Leu), and serine (Ser). Furthermore, using a metabolically inactive control material (tooth dentin) and a control material with normal metabolism (normal articular cartilage), we developed an age adjustment in order to make inferences about the state of protein turnover in cartilage and meniscus.ResultsIn the metabolically inactive control material (n = 25, ages 13 to 80 years) and the normal metabolizing control material (n = 19, ages 17 to 83 years), only Asp + Asn (Asx), Ser, and Leu showed a significant change (increase) in racemization with age (P < 0.01). The age-adjusted proportions of racemized to total amino acid (D/D+L expressed as a percentage of the control material) for Asx, Ser, and Leu when compared with the normal articular cartilage control were 97%, 74%, and 73% in OA meniscal cartilage and 97%, 70%, and 78% in OA articular cartilage. We also observed lower amino acid content in OA articular and meniscal cartilages compared with normal articular cartilage as well as a loss of total amino acids with age in the OA meniscal but not the OA articular cartilage.ConclusionsThese data demonstrate comparable anabolic responses for non-lesioned OA articular cartilage and OA meniscal cartilage but an excess of catabolism over anabolism for the meniscal cartilage.

Highlights

Certain amino acids within proteins have been reported to change from the L form to the D form over time
The age-adjusted proportions of racemized to total amino acid (D/D to total (D+L) expressed as a percentage of the control material) for Asx, Ser, and Leu when compared with the normal articular cartilage control were 97%, 74%, and 73% in OA meniscal cartilage and 97%, 70%, and 78% in OA articular cartilage
We found the relative mean ratios of D to total (D+L) amino acids in our control material to be Asx > Ser > Ala Glx > Leu > Ile (Table 1)

Summary

Introduction

Certain amino acids within proteins have been reported to change from the L form to the D form over time. This process is known as racemization and is most likely to occur in long-lived low-turnover tissues such as normal cartilage. We hypothesized that diseased tissue, as found in an osteoarthritic (OA) joint, would have increased turnover reflected by a decrease in the racemized amino acid content. In proteins with low turnover, this racemization leads to an agedependent increase of D amino acids. All methods of D and L Asp measurement involve high-temperature acid hydrolysis that induces a certain time- and temperaturedependent background racemization; this hampers the ready comparison of results across studies. Successful management of background variability is possible by employing a precise experimental protocol for all samples

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