Biochemistry: Enzyme Regulation, Protein Dynamics, and Protein Study Techniques

Flashcards cover feedback inhibition, allosteric regulation and conformational changes, covalent modifications and signaling proteins, ATP/GTP cycling, protein design, and protein study techniques (homogenate, centrifugation, and chromatography).

What happens during feedback inhibition when methionine accumulates in a biosynthetic pathway?

It blocks the conversion of homoserine into the downstream product, preventing further production.

Name the two internal feedback loops described in the isoleucine biosynthesis example.

One loop involves threonine; the other involves isoleucine itself, both inhibiting further production of isoleucine.

What feedback involving homoserine is described?

Homoserine can feed back to prevent the creation of more homoserine if enough downstream products exist.

Which molecule is mentioned as providing feedback to the original reaction besides methionine and homoserine?

Creatinine.

What does allosteric mean in relation to proteins?

Proteins can adopt two or more conformations; shifts between conformations regulate activity and binding site accessibility.

In the allosteric model, what is the difference between the open and closed conformations regarding substrate binding?

The open form is inactive with no substrate bound; the closed form is the active conformation (ADP-induced) where the active site can bind the substrate (e.g., glucose).

What effect does CTP binding have on the enzyme in the described example?

CTP binds in multiple sites, causing a conformational change that closes the active sites and inhibits activity.

What is phosphorylation?

The addition or removal of phosphate groups, causing conformational changes and altering protein activity.

What does a protein kinase do?

Transfers a phosphate from ATP to a serine (or other amino acid), causing a conformational change and changing activity.

What does a protein phosphatase do?

Removes a phosphate group, reversing the phosphorylation-induced change.

What covalent modifications regulate p53 and what are their effects?

Phosphorylation, acetylation, and ubiquitination; these modifications change p53’s shape and its DNA-binding/transcription activity.

How do regulatory GTP-binding proteins switch on and off?

They are active when bound to GTP and inactive with GDP; hydrolysis to GDP turns them off, and GTP binding reactivates them.

What process uses ATP hydrolysis to drive directed movements of motor proteins?

Motor proteins move along cytoskeletal tracks, using ATP hydrolysis to propel directed movement of cargo or chromosomes.

How does ATP binding and hydrolysis drive the “walk” of a motor protein?

ATP binding induces a conformational change; hydrolysis to ADP+Pi causes a subsequent step; release resets to the starting state for another cycle.

What is a scaffold protein and its function?

A scaffold brings together multiple proteins into a complex by matching domains; the scaffold can be reused after the complex forms.

What is the Rosetta project in protein design?

A computer program used to design proteins from scratch, design sequences, synthesize synthetic genes, and test their function.

What does the video say about the space of possible proteins versus natural proteins?

Nature’s proteins explore a tiny fraction of possible proteins;computational design explores a vastly larger space (e.g., 20^100 possibilities).

Give an example of a therapeutic application mentioned for designed proteins in the video.

Designed protein particles to present viral proteins for stronger immune responses (e.g., vaccines); designs to break down gluten in celiac disease; immune-stimulating cancer therapies.

What is a homogenate in protein studies?

A homogenized mixture containing the contents of broken-open cells, including proteins, lipids, and organelles.

What is centrifugation used for in cell fractionation?

To separate cell components into pellets and supernatants by density and centrifugal force with increasing speeds.

What components are found in the pellet after low-speed centrifugation?

Whole cells, cell nuclei, and the cytoskeleton.

What components are found in the pellet after medium-speed centrifugation?

Mitochondria, lysosomes, and peroxisomes.

What components are found in the pellet after high-speed centrifugation?

Endoplasmic reticulum fragments and other small vesicles.

What is chromatography in this context?

A method to separate components by passing a mixture through a column with a solid matrix and applying solvent.

Name the three types of chromatography mentioned.

Affinity chromatography, ion exchange chromatography, and gel filtration chromatography.

Where would you start if you want to study mitochondria after cell lysis?

Use the pellet from the medium-speed centrifugation step containing mitochondria for further study.

Should you extract proteins from intact cells or after breaking cells?

After breaking the cells to make a homogenate for access to the proteins.

What is the end goal of using these protein study tools?

To access, purify, and characterize proteins and understand their structure and function.