RECOMBINANT IMMUNOGLOBULIN VARIABLE DOMAINS GENERATED FROM SYNTHETIC GENES PROVIDE A SYSTEM FOR IN-VITRO CHARACTERIZATION OF LIGHT-CHAIN AMYLOID PROTEINS

The primary structural features that render human monoclonal light cha ins amyloidogenic are presently unknown. To gain further insight into the physical and biochemical factors that result in the pathologic dep osition of these proteins as amyloid fibrils, we have selected for det ailed study three closely homologous protein products of the light-cha in variable-region single-gene family V kappa IV. Two of these protein s, REC and SMA, formed amyloid fibrils in vivo. The third protein, LEN , was excreted by the patient at levels of 50 g/day with no indication of amyloid deposits. Sequences of amyloidogenic proteins REC and SMA differed from the sequence of the nonpathogenic protein LEN at 14 and 8 amino acid positions, respectively, and these amino acid differences have been analyzed in terms of the three-dimensional structure of the LEN dimer. To provide a replenishable source of these human proteins, we constructed synthetic genes coding for the REC, SMA, and LEN varia ble domains and expressed these genes in Escherichia coli. Immunochemi cal and biophysical comparisons demonstrated that the recombinant V ka ppa IV products have tertiary structural features comparable to those of the patient-derived proteins. This well-defined set of three clinic ally characterized human kappa IV light chains, together with the capa bility to produce these kappa IV proteins recombinantly, provide a sys tem for biophysical and structural comparisons of two different amyloi dogenic light-chain proteins and a nonamyloidogenic protein of the sam e subgroup. This work lays the foundation for future investigations of the structural basis of light-chain amyloidogenicity.