Dimerization of Human Angiogenin and of Variants Involved in Neurodegenerative Diseases

Abstract

Human Angiogenin (hANG, or ANG, 14.1 kDa) promotes vessel formation and is also called RNase 5 because it is included in the pancreatic-type ribonuclease (pt-RNase) super-family. Although low, its ribonucleolytic activity is crucial for angiogenesis in tumor tissues but also in the physiological development of the Central Nervous System (CNS) neuronal progenitors. Nevertheless, some ANG variants are involved in both neurodegenerative Parkinson disease (PD) and Amyotrophic Lateral Sclerosis (ALS). Notably, some pt-RNases acquire new biological functions upon oligomerization. Considering neurodegenerative diseases correlation with massive protein aggregation, we analyzed the aggregation propensity of ANG and of three of its pathogenic variants, namely H13A, S28N, and R121C. We found no massive aggregation, but wt-ANG, as well as S28N and R121C variants, can form an enzymatically active dimer, which is called ANG-D. By contrast, the enzymatically inactive H13A-ANG does not dimerize. Corroborated by a specific cross-linking analysis and by the behavior of H13A-ANG that in turn lacks one of the two His active site residues necessary for pt-RNases to self-associate through the three-dimensional domain swapping (3D-DS), we demonstrate that ANG actually dimerizes through 3D-DS. Then, we deduce by size exclusion chromatography (SEC) and modeling that ANG-D forms through the swapping of ANG N-termini. In light of these novelties, we can expect future investigations to unveil other ANG determinants possibly related with the onset and/or development of neurodegenerative pathologies.

Protective Effects of Recombinant Human Angiogenin in Keratinocytes: New Insights on Oxidative Stress Response Mediated by RNases

“…A further investigation was carried out to verify if it was possible to trace a connection between the above-mentioned stress-induced properties of ANG and its catalytic efficiency. To this purpose, the inactive H13A variant [ 39 , 40 ] was produced to homogeneity, as described in Section 4 . A conformational analysis by CD spectroscopy revealed that H13A_rANG exhibits a similar secondary structure with respect to the parent angiogenin ( Figure 3 A) with a net reduction in the thermal stability ( Figure 3 B).…”

Structure of angiogenin dimer bound to double-stranded RNA

“…While angiogenin is considered to be a primarily monomeric protein, the RNase A family as a whole is known to engage in various homodimeric configurations, including 3D domain-swapped arrangements. Recently, the first evidence of such a 3D-swapped dimer was reported for angiogenin (Fasoli et al, 2021). Evidence was presented for an interaction via the N-terminal helix of angiogenin, but the accompanying homology model shows a different arrangement to the N-terminally mediated interface 2 that is described here.…”

“…Notably, such RNase dimers cannot be bound and inhibited by Rnh1 (Murthy & Sirdeshmukh, 1992). Most recently, dimerization through N-terminal domain swapping has also been reported for angiogenin (Fasoli et al, 2021).…”

Slow Evolution toward “Super-Aggregation” of the Oligomers Formed through the Swapping of RNase A N-Termini: A Wish for Amyloidosis?

“…Hence, we might hypothesize a possible similar behavior for other pt-RNases known to dimerize or oligomerize through the swapping of their N-termini, such as bovine seminal (BS)-RNase [ 50 , 58 ], human RNase 1 variants [ 59 , 60 , 61 , 62 ], human angiogenin [ 63 ], or even amphibian ranpirnase (onconase) [ 64 ].…”