Heat Shock Stress Response on Translation in E. coli

Work done in the lab of Prof. Daniel Wilson at Institute for Biochemistry and Molecular Biology, University of Hamburg, Germany

About author

Haaris Safdari obtained his Bachelor in Biochemistry and Master’s in biotechnology from Aligarh Muslim University, India. He is also a recipient of Sir Syed Global Scholar Award (SSGSA), awarded to meritorious students at university level. He then further carried out his second Masters (Research) from Indian Institute of Science, Bengaluru under Dr. Somnath Dutta, kicking off in the exciting field of structural biology. He is currently pursuing PhD in from Institute for Biochemistry and Molecular Biology, University of Hamburg, Germany under the supervision of Prof. Daniel Wilson. The goal of his research is to understand stress response and pathways in the context of translational machinery by using a combination of biochemical and structural analysis (cryo-electron microscopy).

Interview

How would you explain your research outcomes to the non-scientific community?

In all cells, protein manufacturing is aided by ribosomes, which are situated inside the cell. However, due to some stress such as heat, ribosomes can get stuck (stalling), which can harm cells. We have discovered the conditions that lead to such non-functional ribosomes in Escherichia coli (E. coli), visualized protein called Heat Shock Protein 15 (Hsp15) which is recruited on ribosomes by a technique called cryo-electron microscopy (cryo-EM). We propose a mechanism of Hsp15 protein in the recycling of such non-functional ribosomes.

How do these findings contribute to your research area?

We report a cryo-EM structure of a heat-shock induced protein in E. coli, Hsp15, bound to 50S ribosome subunits (also called large subunit) and peptidyl-tRNA (P-tRNA). Hsp15 was known to bind to such complexes, but there are two new and significant contributions here. First, this report improves on the previous literature by showing that the natural substrate for Hsp15 is 50S subunits bound to peptidyl tRNA arising from the dissociation of 70S subunits during heat shock.

The second contribution is the structure itself, at a much higher resolution than a previously published structure, which it turns out had Hsp15 in the wrong place. Now we see that the E. coli protein binds in the same place as a homolog from B. subtilis, RqcP, which makes sense in terms of their sequence similarity. This finding is important because although RqcP is involved in a ribosome quality control (RQC) pathway in B. subtilis, leading to the alanine-tagging and degradation of the nascent peptide (through the activity of RqcH), that pathway doesn’t seem to exist in E. coli, which lack RqcH. Based on the earlier Hsp15 structure, one might have argued that it differs from RqcP and there is no RQC in E. coli. The new structure establishes that Hsp15 and RqcP similarly bind 50S subunits, arguing that the RQC pathway also exists in E. coli, although we don’t yet know what the downstream factors are.

“This report improves on the previous literature by showing that the natural substrate for Hsp15 is 50S subunits bound to peptidyl tRNA arising from the dissociation of 70S subunits during heat shock.”

What was the exciting moment during your research?

The discovery of an active mechanism in cells that lead to 70S dissociation that gets induced on heat shock was an important and exciting aspect. The results on in vitro purified ribosomes demonstrate that there is no significant 70S dissociation nor 30S degradation. However, the situation inside the cell in vivo is different. It was also remarkable to refute a previous structure of Hsp15 on 50S ribosomes.

What do you hope to do next?

Many times, discoveries in biology lead to new questions and open up a new avenue. The identity of release factor which might release the aberrant nascent chain from the 50S-P-tRNA-Hsp15 complexes remains still unknown but as we state in our manuscript, “Identification of the mystery rescue factor for recycling of 50S-peptidyl-tRNA complexes is one of the exciting perspectives for future study in the field”. We are trying to unravel this aspect and to discover this mystery release factor.

Where do you seek scientific inspiration from?

I seek scientific inspiration from my mentors and major contributors of my research area. I must admit that the book “Gene Machine” by Nobel laureate Prof. Venki Ramakrishnan has left a strong zeal for research in me. Also, I was fortunate enough to learn cryo-electron microscopy in one of the finest labs in India under the supervision of Dr. Somnath Dutta, IISc Bengaluru. My current supervisor, Prof. Daniel Wilson guidance also shaped me to ask important questions in research, redirecting me to address important overall questions in the grand scheme of things.

How do you intend to help Indian science improve?

From what I have learned so far, I try to mentor my colleagues and juniors about the opportunities regarding science in India. I think more practical research experience and internships are needed to be included in the scientific curriculum to instigate the research temperament in the students. As it is wisely said, good science is always collaborative, and we need to improve these to embark on and address important fundamental questions.

Another thing is that we should not consider only Ph.D. as a next career step after completion of Master’s degree just because it seems to be the only option. Unlike master’s courses, which have a defined schedule/timelines to push you up, Ph.D. is a sort of apprenticeship which must only be undertaken if we are passionate about the work, otherwise, it is just a waste of resources and time.

Reference

HA Safdari, S Kasvandik, C Polte, Z Ignatova, T Tenson, DN Wilson (2022) Structure of Escherichia coli heat shock protein Hsp15 in complex with the ribosomal 50S subunit bearing peptidyl-tRNA. Nucleic Acids Research, gkac1035, https://doi.org/10.1093/nar/gkac1035

Copy Editor

Nikita Nimbark

PostGrad in Biotechnology

Nikita Completed her PostGrad in Biotechnology. She has interest in Bioinformatics. Her hobbies include travelling and calligraphy. She is always up for new challenges.

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