ABSTRACT
ATP-dependent proteases of the AAA+ family, including Escherichia coli ClpXP and the eukaryotic proteasome, contribute to maintenance of cellular proteostasis. ClpXP unfolds and translocates substrates into an internal degradation chamber, using cycles of alternating dwell and burst phases. The ClpX motor performs chemical transformations during the dwell and translocates the substrate in increments of 1-4 nm during the burst, but the processes occurring during these phases remain unknown. Here we characterized the complete mechanochemical cycle of ClpXP, showing that ADP release and ATP binding occur nonsequentially during the dwell, whereas ATP hydrolysis and phosphate release occur during the burst. The highly conserved translocating loops within the ClpX pore are optimized to maximize motor power generation, the coupling between chemical and mechanical tasks, and the efficiency of protein processing. Conformational resetting of these loops between consecutive bursts appears to determine ADP release from individual ATPase subunits and the overall duration of the motor's cycle.
Subject(s)
Adenosine Triphosphate/metabolism , Endopeptidase Clp/metabolism , Escherichia coli Proteins/metabolism , Escherichia coli/metabolism , ATPases Associated with Diverse Cellular Activities , Adenosine Diphosphate/metabolism , Adenosine Triphosphatases/chemistry , Adenosine Triphosphatases/metabolism , Endopeptidase Clp/chemistry , Escherichia coli/chemistry , Escherichia coli Proteins/chemistry , Hydrolysis , Kinetics , Models, Molecular , Molecular Chaperones/chemistry , Molecular Chaperones/metabolism , Protein Binding , Protein Conformation , Protein Unfolding , Substrate SpecificityABSTRACT
The purpose of the current study was to analyze the long-term oncologic and functional results and complications associated with limb-sparing surgery and endoprosthetic reconstruction for 23 patients with osteosarcoma of the proximal humerus. There was one Stage IIA lesion, 18 Stage IIB lesions, and four Stage III lesions in this study group. Twenty-two patients were treated with an extraarticular resection that included the deltoid and rotator cuff and one patient was treated with an intraarticular resection that spared the shoulder abductors. In all these patients, the proximal humerus was reconstructed with a cemented endoprosthetic replacement that was stabilized via a technique of static suspension (Dacron tapes) and dynamic suspension (muscle transfers). At latest followup (median, 10 years), 15 patients (65%) were alive without evidence of disease. There were no local recurrences. Prosthetic survival was 100% for the 15 survivors. The Musculoskeletal Tumor Society upper extremity functional score ranged from 24 to 27 (80%-90%). All shoulders were stable and pain-free. Elbow and hand function were preserved in all patients. The most common complication was a transient neurapraxia (n = 8). En bloc extraarticular resection and endoprosthetic reconstruction is a safe and reliable method of limb-sparing surgery for patients with high-grade extracompartmental osteosarcoma of the proximal humerus.
