The folding and aggregation properties of a single KH-domain protein: Ribosome binding factor A (RbfA) from Pseudomonas aeruginosa

Folding Mechanism and Aggregation Propensity of the KH0 Domain of FMRP and Its R138Q Pathological Variant

“…KH0 and KH0-R138Q (50 μM) were pre-heated at 55 °C for 2 h in a water bath. As shown in Figure 4 c,d, the TEM micrographs showed the presence of fibrillar-like aggregates with diameters between 5 and 15 nm, in accordance with previous reports [ 6 , 29 , 38 ]. The fibrils of the KH0 sample appeared dense and curved, with an apparent constant diameter and variable lengths ( Figure 4 c).…”

“…We have shown that the folding of KH0 proceeds through the transient accumulation of an intermediate state, which, according to the global adaptation of the data of the kinetic experiments of rapid (SF) and ultrarapid (TJ) folding, represents an obligatory species along the folding process. This three-state folding mechanism is conserved in KH0-R138Q, as expected for a protein that maintains the same 3D structure [ 11 ] and is reminiscent of the folding mechanism observed for the non-canonical type II KH domain of RbfA [ 6 ]. Furthermore, through complementary biochemical techniques, we have shown that KH0 is prone to form fibrillar-like aggregates and that, even if the folding properties of the KH0 domain appear to be maintained in the R138Q variant, the propensity to aggregate of the variant appears to be increased.…”

“…Moreover, some KH domains defined as non-canonical (or degenerate), lack the conserved GxxG motif in the loop connecting the two alpha-helices of the “core”. Non-canonical KH domains have been identified in both eukaryotic and prokaryotic proteins, such as the bacterial ribosome assembly factor RbfA [ 6 ], the MTH1203 from the archaeon M. thermautotrophicus [ 7 ], the Drosophila P-element somatic inhibitor protein (PSI) [ 8 ], in proteins belonging to the vigilin family [ 9 ], the Rrp40 exosome subunit [ 10 ], and in the fragile X mental retardation protein (FMRP) [ 11 ]. As degenerated KH domains lack the characteristic GxxG loop, they are difficult to predict from sequence analysis and often can be identified only after structure determination; thus, the list of proteins containing this domain will likely increase over time.…”

“…To our knowledge, although there were some preliminary observations [ 29 ], the folding mechanism and aggregation propensity of a KH domain have been studied only recently. In particular, the folding mechanism of RbfA, a KHII-containing protein from P. aeruginosa , was shown to fold according to a three-state mechanism involving a transiently populated intermediate and to form fibrils even in mild conditions [ 6 ].…”

Role of IGF2BPs in head and neck squamous cell carcinoma

“…In this case, the study of folding is helpful to understand the folding properties of KH domain and identify the existence of metastable intermediate states. Whether KH mechanism domain has other functions, such as related to amyloid fiber formation ( 64 ). Further study of the domain is needed to explore more hidden functions of IGF2BPs.…”