| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
Letters to the Editor |
Light Chain Cryoglobulin
1 Endocrine Research Unit, Department of Medicine,2 Biochemistry and, Molecular Biology and, Molecular Pharmacology and, Experimental Therapeutics,3 Division of Hematology and,4 Division of Clinical, Biochemistry and Immunology, Department of Laboratory, Medicine and Pathology, Mayo Clinic College of Medicine, Rochester, MN
aAddress correspondence to this author at: Division of Clinical Biochemistry and Immunology, Department of Laboratory Medicine and Pathology, Hilton 210e, Mayo Clinic, Rochester, MN 55905. Fax 507-266-4088; e-mail abraham.roshini{at}mayo.edu.
To the Editor:
Bence Jones protein with cryoglobulin properties is rare. One of the earliest reports of Bence Jones cryoglobulinuria was made by Alper in 1966 (1) on a patient with multiple myeloma who had a 
Bence Jones cryoglobulin (cryo) in the urine. In this letter, we report a patient with 
Bence Jones cryoglobulinuria. We performed physicochemical analysis of the cryo protein and compared it with a noncryo monoclonal urinary light chain to improve our understanding of the molecular basis of urine cryoglobulin formation.
The patient was a 73-year-old male who had clinical and laboratory findings consistent with multiple myeloma. Serum immunofixation revealed a monoclonal light chain. Urine protein electrophoresis and immunofixation of a randomly collected urine sample revealed the presence of a monoclonal Bence Jones protein. This urine sample formed a gel at 4 °C, which disappeared when the sample was warmed to 37 °C. As a control, we used the urine from a myeloma patient with Bence Jones protein without cryo formation.
We performed single-dimension sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) on the control and cryo urine samples, which revealed a single band at a relative molecular mass (Mr) of 25 000, consistent with monoclonal light chain (Fig. 1
). We then performed gel-filtration chromatography at 4 °C in either "no salt" or "plus NaCl" buffer. Gel-filtration chromatography of the cryo light chain protein in the presence of 100 mmol/L NaCl showed 2 peaks with apparent Mr of 24 000 (29%) and 48 000 (71%). These data indicate that there was a similar distribution of monomeric and dimeric protein species compared with the control light chain; however, the cryo protein was much more compact (spherical) than the control protein in moderate salt solution. In the absence of salt, the cryo protein had an Mr of 68 000, or an apparent mass 2.8-fold larger than that observed in the presence of 100 mmol/L salt. The change in the dimeric form is more dramatic, with an Mr of 280 000, a 5.8-fold increase in apparent mass (Fig. 1
). Because gel-filtration measurements reflect both the mass and shape of the protein, this technique is insufficient of itself to determine whether the cryo light chain has undergone additional polymerization, a large change in conformation, or a combination of both (2)(3). The control Bence Jones protein had 2 molecular species that were consistent with monomer and dimer in the presence or absence of physiologic concentrations of salt.
|
Sedimentation equilibrium experiments were performed on the cryo -light chain protein that had been preequilibrated in the no-salt buffer and run at 6000 and 12 000 rpm at 20 °C (analysis could not be performed at 4 °C because of protein precipitation at higher concentrations). Sedimentation equilibrium centrifugation analysis with a self-association model revealed that the mean molecular mass of the cryo protein in the absence of salt at 20 °C was Mr 60 000. The sedimentation data were consistent with a mixture of 70% dimers and 30% tetramers.
Early studies investigating the mechanisms of temperature- and concentration-dependent cryoprecipitation have shown that low temperatures change the intramolecular environment of the protein, particularly with certain aromatic residues (4). It is possible that there are both salt- and temperature-dependent changes at 20 °C and that only the salt-dependent changes in polymerization are detected by either sedimentation equilibrium or by the apparent Mr measured by gel filtration at 4 °C. Low salt accurately reflects the presence of trimers and hexamers that would form a cryoprecipitate at the higher concentration of light chain protein found in undiluted urine stored at 4 °C. The sodium concentration in urine can vary considerably, and we did not ascertain the sodium in the urine of the cryo patient. It is noteworthy, however, that the control light chain did not show any salt-dependent changes in relative molecular mass, unlike the cryo protein.
It is likely that several physicochemical factors contributed to urine cryoglobulin formation in this patient, including the immunoglobulin light chain protein sequence, resulting from the inherent diversity in the variable regions of immunoglobulins, and the concentration, because the protein had to be diluted substantially to prevent precipitation on the gel-filtration column at 4 °C. In addition, there was no evidence of clinical symptoms related to the urine cryo at physiologic temperature. The urine remained a clear liquid at room temperature and 37 °C and showed complete gel formation only at 4 °C. However, because we do not have data to confirm the role of these various factors, the extent to which these may have contributed to cryo formation in this patient remains speculative.
Footnotes
1 These authors contributed equally to this work. 
References
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
70% and