The amino acid sequence of a human kappa light chain.
ABSTRACT: The amino acid sequence of the light chain of the myeloma protein Dee was studied. The light chain is of the kappa type and of subgroup I. The variable part contains some substitutions that are unique and also some that have been observed already in other kappaI chains (repeated variants). Based on these repeated variants a subdivision of the kappaI subgroup is proposed.
Project description:Glycopeptides have been isolated from tryptic digests of kappa-type light chains separated from human myeloma proteins obtained from the serum of two patients, Car and Rai. The glycopeptides are derived from the variable region of the chain in both cases, but from different sections. On the basis of homology it is deduced that glycopeptide from Car, kappaI type, is derived from position 25-31 whereas that from Rai, kappaII type, is from position 62-77, their sequences being respectively Ala-Ser-Gln-Asn-Ile-Ser and Phe-Ser-Gly-Ser-Gly-Ser-Gly(Thr,Asp)Phe-Thr-Leu-Asx-Ile-Ser-Arg. The significance of the results is discussed in connexion with the nature of the attachment site of carbohydrate to protein.
Project description:Light chain amyloidosis is a devastating protein misfolding disease characterized by the accumulation of amyloid fibrils that causes tissue damage and organ failure. These fibrils are composed of monoclonal light chain protein secreted from an abnormal proliferation of bone marrow plasma cells. We previously reported that amyloidogenic light chain protein AL-09 adopts an altered dimer while its germline protein (kappaI O18/O8) forms a canonical dimer observed in other light chain crystal structures. In solution, conformational heterogeneity obscures all NMR signals at the AL-09 and kappaI O18/O8 dimer interfaces, so we solved the nuclear magnetic resonance structure of two related mutants. AL-09 H87Y adopts the normal dimer interface, but the kappaI Y87H solution structure presents an altered interface rotated 180 degrees relative to the canonical dimer interface and 90 degrees from the AL-09 arrangement. Our results suggest that promiscuity in the light chain dimer interface may promote new intermolecular contacts that may contribute to amyloid fibril structure.
Project description:That structural abnormalities may be responsible for nonamyloid immunoglobulin (Ig) light chain deposition disease (LCDD) is suggested by previous results of Ig biosynthesis studies, but this hypothesis was not documented at the molecular level. We report on the first complete primary sequence deduced from cDNA analysis of a kappa light chain responsible for LCDD associated with an apparently nonsecretory myeloma. Bone marrow myeloma cells contained intracellular kappa chains and no heavy chains by immunofluorescence. Kidney biopsy showed typical nonamyloid PAS-positive kappa chain deposits. SDS-PAGE analysis of material extracted from a kidney biopsy specimen and of Ig produced by the myeloma cells revealed kappa chains of abnormally high apparent molecular mass (30,000). Comparison of the NH2-terminal aminoacid sequence of the kappa chain deposited in the kidney and of the complete sequence of several identical kappa cDNA clones from bone marrow cells showed the identity of the tissue deposited and plasma cell kappa chain. The kappa mRNA had an overall normal structure and corresponded to the V kappa IV gene rearranged to J kappa 1 and followed by a normal constant exon of the Km(3) allotype. The variable sequence differed from the V kappa IV germline gene by nine point mutations, including an Asp----Asn substitution at position +70 resulting in a potential N-glycosylation site. In vitro biosynthesis experiments and treatment with N-glycosidase provided evidence for the intracellular glycosylation of the monoclonal kappa chain. The peculiar sequence and the glycosylation of a kappa chain of the rare V kappa IV subgroup might be responsible for structural abnormalities leading to tissue deposition.
Project description:Amyloidoses are devastating and currently incurable diseases in which the process of amyloid formation causes fatal cellular and organ damage. The molecular mechanisms underlying amyloidoses are not well known. In this study, we address the structural basis of immunoglobulin light chain amyloidosis, which results from deposition of light chains produced by clonal plasma cells. We compare light chain amyloidosis protein AL-09 to its wild-type counterpart, the kappaI O18/O8 light chain germline. Crystallographic studies indicate that both proteins form dimers. However, AL-09 has an altered dimer interface that is rotated 90 degrees from the kappaI O18/O8 dimer interface. The three non-conservative mutations in AL-09 are located within the dimer interface, consistent with their role in the decreased stability of this amyloidogenic protein. Moreover, AL-09 forms amyloid fibrils more quickly than kappaI O18/O8 in vitro. These results support the notion that the increased stability of the monomer and delayed fibril formation, together with a properly formed dimer, may be protective against amyloidogenesis. This could open a new direction into rational drug design for amyloidogenic proteins.
Project description:1. The specificity of combination of heavy and light chains from selected human immunoglobulins was examined in the light of greater structural information than in previous studies. Heavy (gamma) chains from immunoglobulin G (kappa) myeloma proteins were allowed to combine with their homologous light (kappa) chains or with other kappa chains of the same variable-region subgroup. The affinity of each such pairing was assessed by having the test kappa chain compete with a standard population of normal light chains. 2. There was a spread of affinities among the heavy-light pairings with the homologous pairings having an average affinity significantly higher than the heterologous pairings. 3. It follows that (a) the preference shown for homologous heavy-light pairings is not explicable simply in terms of the known subdivisions of the variable and constant regions of the chains, and (b) it is unlikely that those residues specifying the subgroups of kappa-chain variable regions have a predominant role in the formation of interchain bonds with the gamma-chain variable regions.
Project description:Mechanisms of amyloidogenesis are not well understood, including potential structural contributions of mutations in the process. Our previous research indicated that the dimer interface of amyloidogenic immunoglobulin light chain protein AL-09 is twisted 90 degrees relative to the protein from its germline sequence, kappaI O18/O8. Here we report a systematic restoration of AL-09 to its germline sequence by mutating the non-conservative somatic mutations located in the light chain dimer interface. Among these mutants, we find a correlation between increased thermodynamic stability and an increase in the lag time for fibril formation. The restorative mutant AL-09 H87Y completes the trifecta and restores the dimer interface observed in kappaI O18/O8, emphasizing the potential importance of the structural integrity of these proteins to protect against amyloidogenicity. We also find that adding amyloidogenic mutations into the germline protein illustrates mutational cooperativity in promoting amyloidogenesis.
Project description:MPC 11 mouse myeloma cells synthesize two immunoglobulin kappa light chains, coded by two separate genes. One of these Kappa-chains has no variable region and is degraded intracellularly. The other is a full-length kappa-chain contaning both variable and constant regions: this chain is secreted, both by itself and combined with heavy chains in molecules of immunoglobulin G. This paper reports the amino acid sequence of the myeloma MPC 11 full-length kappa-chain. The chain is unusual in having 12 extra residues at its N-terminus when its sequence is aligned with those of other mouse kappa-chains; no other anomalies were found in its sequence.
Project description:1. The light chains of human immunoglobulin (Ig) exist in two forms, kappa (type K) and lambda (type L). The two types of chains can be partially separated by taking advantage of the fact that lambda-chains, for the most part, dissociate from reduced Ig at higher pH than do the kappa-chains. The same difference in dissociation of type K and L chains was observed with myeloma IgG and IgA proteins, but not with pathological IgM proteins. 2. When analysed in urea-glycine starch gels, pH7, both kappa- and lambda-chains show ten electrophoretic bands having the same mobilities as those of the whole light-chain subfractions. Normal kappa- and lambda-chains show similar differences in overall amino acid composition to those previously found with myeloma kappa- and lambda-chains and type K and L Bence-Jones proteins.
Project description:Although there are many known Mendelian genes linked to epileptic or developmental and epileptic encephalopathy (EE/DEE), its genetic architecture is not fully explained. Here, we address this incompleteness by analyzing exomes of 743 EE/DEE cases and 2366 controls. We observe that damaging ultra-rare variants (dURVs) unique to an individual are significantly overrepresented in EE/DEE, both in known EE/DEE genes and the other non-EE/DEE genes. Importantly, enrichment of dURVs in non-EE/DEE genes is significant, even in the subset of cases with diagnostic dURVs (P = 0.000215), suggesting oligogenic contribution of non-EE/DEE gene dURVs. Gene-based analysis identifies exome-wide significant (P = 2.04 × 10-6) enrichment of damaging de novo mutations in NF1, a gene primarily linked to neurofibromatosis, in infantile spasm. Together with accumulating evidence for roles of oligogenic or modifier variants in severe neurodevelopmental disorders, our results highlight genetic complexity in EE/DEE, and indicate that EE/DEE is not an aggregate of simple Mendelian disorders.
Project description:Bence Jones proteinuria is a disorder that is defined by the excretion of monoclonal light-chain protein. About 15-20% of patients with multiple myeloma secrete monoclonal light chains only, without expression of the normal immunoglobulin heavy chain, which constitutes light-chain multiple myeloma. The definition, prevalence, and progression of these premalignant phases of light-chain multiple myeloma have not been fully characterised. We aimed to identify a subset of patients with idiopathic Bence Jones proteinuria who had a high risk of progression to light-chain multiple myeloma analogous to that seen in patients with smouldering multiple myeloma.In this retrospective cohort study, we studied all patients seen at the Mayo Clinic (Rochester, MN, USA) within 30 days of diagnosis of idiopathic Bence Jones proteinuria between Jan 1, 1960, and June 30, 2004. Inclusion criteria were monoclonal light chain in the urine (?0·2 g/24 h), absence of intact monoclonal immunoglobulin (M protein) in the serum, and no evidence of multiple myeloma, light-chain amyloidosis, or other related plasma-cell proliferative disorders. The primary endpoint was progression to symptomatic multiple myeloma or light-chain amyloidosis. We examined the cumulative probability of progression and the association of potential risk factors on progression rates to identify patients with a high risk of progression to multiple myeloma or light-chain amyloidosis.We identified 101 patients with idiopathic Bence Jones proteinuria. During 901 total person-years of follow-up, 27 (27%) patients developed multiple myeloma and seven (7%) developed light-chain amyloidosis. The major risk factors for progression were amount of urinary excretion of M protein per 24 h, proportion of bone marrow plasma cells, presence of a markedly abnormal free-light-chain ratio (<0·01 or >100), and reduction of all three uninvolved immunoglobulins. Based on the risk of progression, monoclonal light-chain excretion of 0·5 g/24 h or greater or at least 10% bone marrow plasma cells, or both, in the absence of end-organ damage was used to define light-chain smouldering multiple myeloma. The cumulative probability of progression to active multiple myeloma or light-chain amyloidosis in patients with light-chain smouldering multiple myeloma was 27·8% (95% CI 14·2-39·2) at 5 years, 44·6% (27·9-57·4) at 10 years, and 56·5% (36·3-70·2) at 15 years.Light-chain smouldering multiple myeloma as defined in this study is associated with a high risk of progression to symptomatic light-chain multiple myeloma, and this subset of patients needs careful observation and could benefit from clinical trials of early intervention.Jabbs Foundation (Birmingham, UK), US National Cancer Institute, and Henry J Predolin Foundation (Madison, WI, USA).