Angiogenin (Ang) and the Ribonuclease A Family

Flashcards covering the structure, function, evolution, and medical relevance of human Angiogenin based on lecture materials.

What is the full name of the enzyme abbreviated as Ang?

Angiogenin

In what year was human angiogenin discovered?

$$1985$$

Who discovered angiogenin at Harvard University?

Bert Vallee and colleagues

From what medium was angiogenin first isolated?

Human tumour cell-conditioned medium

How many residues are in the single-chain polypeptide of angiogenin?

$123$ residues

Angiogenin is a homologue of which well-characterized enzyme family?

The Ribonuclease A (RNase A) family

What is the definition of angiogenesis?

The growth of new blood vessels in both normal tissue physiology and pathological settings.

Insufficient blood vessel growth can lead to which three medical conditions?

Ulcers, strokes, and wounds (defective healing)

Excessive blood vessel proliferation favors which three pathological states?

Tumour growth and spreading, diabetic retinopathy, and arthritis

What is the term for new blood vessel growth originating from endothelial cell precursors (angioblasts)?

Vasculogenesis

Angiogenin is also known by what family name and number?

RNase 5

How does the ribonucleolytic activity of angiogenin compare to that of RNase A?

It is $10^5$ to $10^6$ times weaker than that of RNase A.

True or False: RNase A is an angiogenic molecule.

False

What is the sequence identity percentage between angiogenin and RNase A?

$$33\%$$

When considering conserved residues, what is the similarity percentage between Ang and RNase A?

$$46\%$$

Name four primary physiological roles for angiogenin.

Wound healing, placental development, establishment of pregnancy, and innate immunity.

In fetal RNA, at which three marks of gestation was angiogenin expression noted?

$19$ weeks, $9$ weeks, and $6$ weeks

Name five fetal tissues listed that express human angiogenin.

Lung, heart, brain, skin epithelium, and melanocytes.

Which adult tissues express human angiogenin according to the transcript?

Prostate, brain, colon, lung, kidney, and retina.

In which two types of cancers is angiogenin highly expressed according to NCBI data?

Germ cell and testicular tumours.

What four aspects of angiogenin are required for the process of angiogenesis?

Ribonucleolytic activity, stimulation of basement membrane degradation, movement/stimulation of signal transduction, and nuclear translocation.

By what mechanism is angiogenin internalized into cells for nuclear translocation?

Receptor mediated endocytosis

When was the three-dimensional structure of human angiogenin determined?

In the early $1990s$

How many disulfide bridges does RNase A have compared to angiogenin?

RNase A has four, while angiogenin has three.

What three residues constitute the catalytic triad in angiogenin?

$\text{His-13}$, $\text{Lys-40}$, and $\text{His-114}$

Which specific residue acts as a 'tollgate' blocking the active site of angiogenin?

Glutamine 117 ($\text{Gln-117}$)

What must the angiogenin molecule undergo to bind and cleave RNA?

A conformational change

Replacing which catalytic residues abolishes the angiogenic activity of angiogenin?

$\text{His-13}$, $\text{Lys-40}$, and $\text{His-114}$

Mutation of which substrate-binding residue reduces angiogenic activity?

$$\text{Thr-44}$$

What is the biological effect of replacing $\text{Asp-116}$ with $\text{His}$ in angiogenin?

It increases both ribonucleolytic and angiogenic activities.

Crystal structures of which two variants proved that loss of function in Ang was chemical and not structural unfolding?

$\text{H13A}$ and $\text{K40Q}$

In which specific sub-cellular location does angiogenin accumulate to stimulate pre-rRNA production?

The nucleolus

Angiogenin antagonists prevent how many types of subcutaneous tumours in mice?

$7$ types

In mouse models, angiogenin antagonists prevent the metastatic spread of which two cancers?

Prostate and breast cancer

Name four types of human cancers where serum angiogenin levels are elevated.

Pancreatic, colon, gastric, and cervical cancers.

What condition upregulates angiogenin expression in metastatic human melanoma?

Hypoxia

What is the name of the murine monoclonal antibody (mAb) that prevents human tumour appearance?

$$\text{mAb 26-2F}$$

What is the binding affinity ($\text{IC}_{50}$) of $\text{mAb 26-2F}$?

$$1.6\,\text{nM}$$

At what resolution was the crystal structure of the angiogenin bound to $\text{Fab 26-2F}$ solved?

$$2.0\,\text{\AA}$$

What is the full name of the protein abbreviated as hRI?

Human placental Ribonuclease Inhibitor (or RNH1)

What is the molecular size of hRI?

$$50\,\text{kDa}$$

hRI is a leucine-rich protein that binds to RNase A and Ang with a $Ki$ of approximately what?

$\text{Ki} = 0.5\,\text{fM}$ for Angiogenin and $\text{Ki} = 40\,\text{fM}$ for RNase A.

What are the three main limitations of hRI as a therapeutic drug?

Broad specificity (binds many RNases), lability (sensitive to oxidation), and large size.

What is the stoichiometry of the hRI-Angiogenin complex?

$$1:1$$

What small-molecule inhibitor from NCI showed reduction in prostate cancer tumor progression?

$\text{NCI compound 65828}$ (or $\text{N-65828}$)

The ANG gene was the first loss-of-function gene identified in which neurodegenerative disease?

Amyotrophic lateral sclerosis (ALS)

Besides ALS, what other neurodegenerative disease is angiogenin implicated in?

Parkinson\u2019s disease

Where is angiogenin strongly expressed within the developing central nervous system?

Differentiating and adult motor neurons

In which neuronal structure does angiogenin localize?

Growth cones

Angiogenin co-localizes with what specific neuronal fibers?

Neurofilaments

Angiogenin is important for the assembly of what cellular compartments during stress?

Stress granules (SG)

True or False: The weak RNase activity of Ang is essential for inducing stress granules.

True

What is the recently identified cell surface receptor for angiogenin?

Plexin B2

Plexins are generally cell surface receptors for which proteins?

Semaphorins (Sema)

Sema-Plexin interactions are involved in what four biological processes?

Neurogenesis, angiogenesis, tumour progression, and immune responses.

In stress responses, angiogenin cleaves tRNAs at which location?

The anticodon loop

What is the secondary consequence of angiogenin cleaving tRNA during stress?

Translation repression

According to Cryo-EM, what cellular component acts as the activator of angiogenin?

The cytosolic ribosome

In the cryo-EM structure, where is angiogenin bound on the ribosome?

In the A site of the $80\text{S}$ ribosome.

Which part of the angiogenin molecule is rearranged by interaction with the ribosome to activate the catalytic center?

The C-terminal tail

How many orders of magnitude more efficient is tRNA cleavage when Ang is bound to the ribosome?

Several orders of magnitude

What component acts as the 'specificity factor' for the tRNA substrate in the ribosome-Ang complex?

The ribosome

Ribosomes with what type of site are more abundant during cellular stress to activate Ang?

A vacant A site

Angiogenin produced by Paneth cells in the intestine has what function?

Anti-microbial activity

Angiogenin is important in preventing the degranulation of which immune cells?

Polymorpho-nuclear leukocytes

What happens to the serum levels of angiogenin during inflammation?

They are elevated

What is the $Ki$ for the RNase A - pdUppA-3'-p complex?

$$\text{Ki} = 27\,\text{nM}$$

Which type of nuclease catalyzes the degradation of RNA into smaller components and is the model for Ang?

Ribonuclease A (RNase A)

What is the basic shape of the RNase A molecule in 3D?

Kidney-shaped

What kind of protein is RNase A in terms of secondary structure composition?

An alpha-beta protein ($\alpha$-helices and $\beta$-sheets)

Based on the transcript, what is the role of the B1 pyrimidine site in angiogenin?

It is blocked by residue $\text{Gln-117}$.

What specific molecules are produced when Ang cleaves tRNAs?

$$\text{tiRNA}$$

What was the purpose of creating peptide mimetics based on the $26\text{-}2\text{F}$ antibody crystal structure?

For use in anti-cancer therapy.

Which specific human cell line was used to test N-65828 in mice?

$\text{PC-3}$ prostate tumour cells

Is the loss of function in Ang-ALS variants always due to decreased RNase activity?

No, variants that either decrease or increase the RNase activity can affect neuronal survival.