Biology subjects

Bhowmick, J.

Publications and source records attributed to Bhowmick, J..

2 recordsLinked to original sources

bioRxiv2026

Structural insights into the inhibition of sickle hemoglobin polymerization by asymmetric hemoglobin tetramer HbFS (α2γβS)

Sickle cell disease (SCD) is caused by a single amino acid substitution in the {beta}S globin chain at 6th position (6E[->]V). This results in polymerization of deoxy state of sickle hemoglobin (HbS), followed by its precipitation and subsequent sickling of erythrocytes. These deformed cells can block small capillaries (vaso-occlusion), causing cardiovascular complications, ultimately leading to ischemia-reperfusion injury, severe oxygen deficiency, and progressive systemic damage. Occasionally, patients with SCD have been observed to produce exorbitantly high levels of fetal hemoglobin (HbF), which has been linked with the inhibition of HbS polymerization. One of the effects of hydroxyurea, the most commonly used therapeutic for SCD, is to elevate HbF levels. However, the mechanism of inhibitory role of HbF on HbS polymerization is largely unknown. This study attempts to gain insights into the mechanisms involved in this process by means of native mass spectrometry, ion mobility mass spectrometry, and hydrogen deuterium exchange-based mass spectrometry (H/DX-MS). The conformational flexibility of asymmetric hemoglobin, HbFS (2{gamma}{beta}S), for the observed regions in the tetrameric molecule appears to be more in the deoxy state as compared to the oxy state, eventually leading to reduced polymerization of sickle hemoglobin in patients with SCD that express elevated HbF levels.

biochemistry↗

bioRxiv2021

Mechanistic insights into global suppressors of protein folding defects

Most amino acid substitutions in a protein either lead to partial loss of function or are near neutral. Several studies have shown the existence of second-site mutations that can rescue defects caused by diverse loss of function mutations. Such global suppressor mutations are key drivers of protein evolution. However, the mechanisms responsible for such suppression remain poorly understood. To address this, we characterized multiple suppressor mutations both in isolation and in combination with inactive mutants. We examined six global suppressors of the bacterial toxin CcdB, the known M182T global suppressor of TEM-1 {beta}-lactamase, the N239Y global suppressor of p53-DBD and three suppressors of the SARS-CoV-2 spike Receptor Binding Domain. The suppressors both alone, and in conjunction with inactive mutants, stabilise the protein both thermodynamically and kinetically in-vitro, predominantly through acceleration of the refolding rate. When coupled to inactive mutants they promote increased in-vivo solubilities as well as regain-of-function phenotypes. In the case of CcdB, where novel suppressors were isolated, we determined the crystal structures of three such suppressors to obtain insight into the specific molecular interactions responsible for the observed effects. While most individual suppressors result in small stability enhancements relative to wildtype, which can be combined to yield significant stability increases, thermodynamic stabilisation is neither necessary nor sufficient for suppressor action.

biophysics↗