Glycine structure is the simplest in biochemistry: a single hydrogen atom where every other standard amino acid carries a side chain. Proline structure is the strangest: its side chain loops back and bonds to its own backbone nitrogen. Between them, these two residues decide where a peptide chain can bend, where it cannot, and why a short synthetic peptide can look like two compounds on an HPLC trace when it is only one.
This guide draws each structure, explains why glycine is achiral and why proline is not really an “imino acid”, covers proline’s slow cis-trans switching, and applies it to Semax, Selank and GHK.
The short answer
- Glycine is the smallest amino acid. Its formula is C2H5NO2 (75.07 g/mol), and its side chain is a single hydrogen atom, so its alpha carbon carries two identical hydrogens.
- Glycine is achiral. With two identical substituents on the alpha carbon, glycine has no stereocentre and no L or D form; it is the only one of the 20 standard amino acids without one.
- Proline is a cyclic secondary amine. Its three-carbon side chain closes onto the backbone nitrogen to form a five-membered pyrrolidine ring; its formula is C5H9NO2 (115.13 g/mol).
- “Imino acid” is an obsolescent label for proline. In IUPAC usage an imino acid carries a C=N group; proline has none, and IUPAC marks the older sense, which used proline as its example, as obsolescent.
- Proline locks the backbone and removes a hydrogen-bond donor. The ring fixes the phi angle and, inside a chain, leaves proline’s nitrogen with no hydrogen, which is why proline disrupts alpha helices; glycine disrupts them too, through excess flexibility.
- Bonds in front of proline switch slowly between cis and trans. About 5% of X-Pro bonds in folded proteins are cis, against a few hundredths of a percent for other peptide bonds; the switch takes long enough to show up in chromatography.
Glycine and proline at a glance
Gly · G · aminoacetic acid | Pro · P · pyrrolidine-2-carboxylic acid
- Glycine formula
- C2H5NO2, 75.07 g/mol; side chain –H
- Proline formula
- C5H9NO2, 115.13 g/mol; side chain –CH2CH2CH2– bonded back to N
- Chirality
- Glycine: none. Proline: one stereocentre; proteins use L-proline
- pKa values
- Glycine 2.34 and 9.60; L-proline 1.99 and 10.60 (PubChem, citing the Merck Index)
- Residue mass in a chain
- Gly 57.02 Da, Pro 97.05 Da (monoisotopic, calculated)
- Backbone N–H in a chain
- Glycine: yes. Proline: no
- Cis X-Pro bonds
- About 5.2% in a survey of folded protein structures
Where the evidence stands
- Structures and constants
- Conformation surveys
- Isomerisation kinetics measured
- Predicted for any given peptide
The chemistry is textbook-settled. What is not settled is how much cis proline a particular short peptide carries under particular conditions; that has to be measured.
Glycine structure: the amino acid with no side chain
Every standard amino acid has an alpha carbon bonded to an amino group, a carboxylic acid group, a hydrogen atom and a side chain, or R group. Glycine’s R group is a second hydrogen atom. Its condensed formula is H2N–CH2–COOH (C2H5NO2, 75.07 g/mol). Our amino acid codes guide lists it as Gly, or G.
Why glycine is achiral
A carbon atom is a stereocentre when it carries four different groups, because only then does it have a mirror image that cannot be superimposed on it. Glycine’s alpha carbon carries two identical hydrogens, so its mirror image is the same molecule. Glycine therefore has no L and D forms and does not rotate plane-polarised light. Every other standard amino acid has L and D forms; our guide to D-amino acids explains why that matters for peptide design.
The zwitterion and its two pKa values
In water near neutral pH, glycine exists mainly as a zwitterion, +H3N–CH2–COO−: the carboxyl group has given up a proton and the amino group has taken one. PubChem’s record, citing the Merck Index, gives pKa values of 2.34 for the carboxyl group and 9.60 for the amino group. Inside a peptide chain, glycine contributes a residue of C2H3NO, 57.02 Da monoisotopic, the number used when checking a sequence against a mass spectrum.
Proline structure: a ring that closes onto the backbone
Proline’s side chain is three methylene groups, –CH2CH2CH2–, but instead of ending freely the chain bonds back to the alpha-amino nitrogen. The result is a five-membered pyrrolidine ring that contains the nitrogen, the alpha carbon and the three side-chain carbons. Its systematic name is pyrrolidine-2-carboxylic acid, its formula is C5H9NO2, and its molar mass is 115.13 g/mol. The alpha carbon is still a stereocentre, and proteins use L-proline. PubChem gives L-proline pKa values of 1.99 and 10.60.
Secondary amine, not imino acid
Because the nitrogen is bonded to two carbons, free proline is a secondary amine; all the other standard amino acids are primary amines. Older textbooks call proline an “imino acid”, and the label persists online. It is a misnomer by current conventions. The IUPAC glossary of class names defines imino acids as carboxylic acids bearing an imino group, HN=, a carbon-nitrogen double bond, which proline does not have. The same entry records the older sense, cyclic aza-acids “e.g. proline”, and marks it obsolescent. The accurate description is a cyclic secondary amino acid.
A common error on page one. Some popular explainer pages say proline has “no alpha hydrogen”. It does. What proline lacks, once it is inside a peptide chain, is a hydrogen on its backbone nitrogen, because that nitrogen’s third bond goes to the ring.
What glycine and proline do to a peptide backbone
A peptide backbone has two rotatable bonds per residue, the phi and psi angles; the peptide bond itself, omega, is planar. Side chains limit which phi and psi combinations a residue can adopt. Glycine and proline sit at the two extremes.
| Property | Glycine | Proline |
|---|---|---|
| Side chain | –H | Ring bonded to backbone N |
| Backbone freedom | Unusually wide; reaches regions other residues avoid, including the left-handed region | Phi fixed by the ring; psi still varies |
| Backbone N–H in a chain | Present | Absent (tertiary amide) |
| Effect inside an alpha helix | Destabilising: entropy cost of freezing a flexible residue | Disruptive: no N–H donor and a fixed phi angle |
| Bond in front of it | Strongly trans, like most peptide bonds | Cis and trans both populated; slow exchange |
| Where it stands out | Turns, loops, tight packing (collagen) | Polyproline II stretches, collagen |
Glycine: flexibility as a cost
Ho and Brasseur’s 2005 analysis of glycine Ramachandran plots described five distinct populated regions, including a left-handed helical region that bulkier residues rarely occupy. That freedom is useful in tight turns and loops. It also helps explain why glycine is a poor helix former: the usual reasoning is that fixing a very flexible residue into a helix costs more conformational entropy than fixing a constrained one. Chakrabartty, Schellman and Baldwin measured the size of the effect in a 17-residue alanine-lysine model peptide and reported a helix-propensity ratio of about 100 between alanine and glycine.
Proline: rigidity and a missing donor
Proline’s ring holds the phi angle in a narrow band of negative values; in the polyproline II conformation it sits near −75°. Inside a chain, the nitrogen forms its peptide bond and its ring bond, so no hydrogen is left on it. In an alpha helix, each backbone N–H donates a hydrogen bond to the carbonyl four residues earlier, so a proline in the middle of a helix leaves a gap in that pattern, and its fixed phi angle limits how the chain can adjust around the gap. Ho and Brasseur also showed that proline constrains the residue in front of it, the “pre-proline” position, which has a Ramachandran plot of its own.
Proline cis-trans isomerization
A peptide bond has partial double-bond character, so it is planar and comes in two forms: trans, with consecutive alpha carbons on opposite sides, and cis, with them on the same side. For an ordinary peptide bond the trans form is so strongly favoured that cis bonds are rare. A 2013 review by Craveur and colleagues, summarising the survey of 571 non-redundant protein structures by Jabs, Weiss and Hilgenfeld, reports cis conformations at 5.2% of X-Pro bonds but only 0.03% of bonds in front of any other residue. In random-coil polypeptides, the same review gives cis fractions for X-Pro bonds of 5% to 30%, depending mainly on the neighbouring residues.
The usual structural explanation is steric. In front of an ordinary residue, the cis form crowds two alpha carbons together, while the trans form does not. In front of proline, the nitrogen carries the ring’s delta carbon in place of a hydrogen, so both forms involve some crowding and the energy gap between them narrows.
Why the switch is slow
Rotating about a bond with partial double-bond character requires passing a high energy barrier; dipeptide simulations summarised in the same review put it at about 20 kcal/mol. Interconversion is therefore slow enough to be seen on the timescale of protein refolding and of a chromatographic run. In 1975, Brandts, Halvorson and Brennan proposed that this explained the slow phase seen in protein refolding experiments, and in 1987 Lang, Schmid and Fischer showed that an enzyme, prolyl isomerase, catalyses protein folding by speeding the step.
Short synthetic peptides are a special case
Much early cis-proline data came from short peptides. Alderson and colleagues, using NMR on intact unfolded proteins, found cis-proline populations below 5% at every X-Pro bond of alpha-synuclein, and showed that short peptides of the same sequence overestimate the cis population, especially when their termini are free and oppositely charged. A heptapeptide with free ends, like many research peptides, is that kind of molecule, so a measurable cis population should be expected.
Semax and Selank, both ending in Pro-Gly-Pro, are stocked as lyophilized research reagents, priced in CAD.
Browse cognitive & neuroWhat it looks like on an HPLC trace
Because cis and trans conformers interconvert slowly, a chromatography column can partly separate them. Nishikawa and colleagues ran X-Pro dipeptides such as Leu-Pro by reversed-phase liquid chromatography and saw two peaks, one for each conformer, at low column temperature. As temperature or retention time increased, interconversion during the run blurred the peaks into a single broad one. For a proline-containing peptide, a doubled or shouldered main peak that merges when the column is warmed points toward conformers rather than an impurity. Mass spectrometry is the decisive check: conformers have identical mass, while a genuine impurity usually does not. See how to read an HPLC chromatogram for peak shape, and the HPLC testing guide for how purity is calculated from peak area.
Glycine and proline together: collagen and the polyproline II helix
The clearest example of the two working together is collagen. Its strands follow a repeating Gly-Xaa-Yaa pattern in which glycine occupies every third position, because the three strands pack so tightly around the shared axis that only a residue with no side chain fits at the centre. Proline is common at the Xaa position and 4-hydroxyproline at Yaa, and Jenkins and Raines describe Gly-Pro-Hyp as the most stabilising triplet in natural collagen. Each strand on its own adopts a left-handed polyproline II-type helix, and three of them wind together into the triple helix.
The polyproline II (PPII) helix is not limited to collagen. Surveying 274 protein chains, Stapley and Creamer found short PPII stretches in most proteins, with proline the most common residue. They reported phi and psi angles of about −75° and 145° and a threefold rotational symmetry, which makes PPII an extended, left-handed helix with three residues per turn, quite unlike the compact alpha helix.
Glycine and proline in real peptide sequences
Sequences below are from Pure Peptides Canada’s product pages, in the one-letter code explained in our sequence notation guide:
- Semax, MEHFPGP. ACTH(4-7) followed by a Pro-Gly-Pro tail. It contains two X-Pro bonds, Phe-Pro and Gly-Pro, so in solution it can carry up to four cis/trans combinations.
- Selank, TKPRPGP. Tuftsin (TKPR) followed by the same tail. It contains three prolines and three X-Pro bonds, Lys-Pro, Arg-Pro and Gly-Pro, giving up to eight combinations.
- GHK, Gly-His-Lys. The tripeptide begins with glycine, so its N-terminal residue is achiral and only histidine and lysine carry stereocentres. It contains no proline.
The Pro-Gly-Pro extension was added to slow enzymatic breakdown, as our Semax vs Selank comparison explains. For an analyst, the same tail is a reason to confirm any split peak by mass before calling it an impurity. Our overview of dipeptides, tripeptides and oligopeptides places GHK and other short sequences in context, and what are peptides covers the peptide bond itself.
Counting is not predicting. These combination counts are upper limits. How much of each conformer a peptide actually carries depends on sequence, solvent, pH and temperature, and has to be measured, typically by NMR.
What’s established and what isn’t
Reasonably well established
- Glycine’s side chain is a hydrogen atom, so glycine is achiral.
- Proline is a cyclic secondary amine; “imino acid” is an obsolescent IUPAC label for it.
- Inside a chain, proline has no backbone N–H and a constrained phi angle.
- X-Pro bonds are cis far more often than other peptide bonds, and interconvert slowly.
- Collagen requires glycine at every third position.
Open questions
- The cis fraction for any specific short peptide, which varies with neighbours, termini and conditions.
- How closely conformer data from short model peptides reflect longer chains; Alderson’s work shows they can overestimate cis populations.
- How far proline-rich heptapeptides such as Selank adopt PPII in solution; the sources reviewed here do not say.
Sourcing glycine- and proline-rich peptides in Canada
For a lab buying research peptides in Canada, the structure points above become documentation questions.
- Ask for identity by mass, not only purity by HPLC. A proline-rich peptide can show conformer peaks; a mass-spectrometry result confirms what the main peak is. See how to read a peptide mass spectrum.
- Check the certificate matches your vial. The batch number on the report should match the batch on the label. Our COA guide lists what else a useful certificate contains.
- Store lyophilized material cold, dry and dark; see the storage guide.
SemaxMEHFPGP: two X-Pro bonds in a Pro-Gly-Pro tailView product
SelankTKPRPGP: three prolines and three X-Pro bondsView product
GHK-CuGly-His-Lys copper complex: an achiral glycine at the N-terminusView product
Research use only. Pure Peptides Canada products are laboratory reagents for in-vitro research, not for human or veterinary use. This article explains published research for education; it is not medical, dietary or dosing guidance.
Frequently asked questions
What is the structure of glycine?
Glycine is H2N-CH2-COOH, molecular formula C2H5NO2 and molar mass 75.07 g/mol. Its alpha carbon carries an amino group, a carboxylic acid group and two hydrogen atoms, because its side chain is a single hydrogen. Near neutral pH it exists mainly as the zwitterion +H3N-CH2-COO-.
Why is glycine not chiral?
A chiral carbon needs four different groups attached to it. Glycine’s alpha carbon carries two identical hydrogen atoms, so the molecule is identical to its mirror image. That makes glycine the only one of the 20 standard amino acids with no L and D forms.
What is the structure of proline?
Proline is pyrrolidine-2-carboxylic acid, molecular formula C5H9NO2 and molar mass 115.13 g/mol. Its three-carbon side chain bonds back to the backbone nitrogen, forming a five-membered ring. This makes free proline a secondary amine, and once it is inside a peptide chain its nitrogen carries no hydrogen.
Is proline an imino acid?
Not by current IUPAC conventions. IUPAC defines imino acids as carboxylic acids bearing an imino group, HN=, which contains a carbon-nitrogen double bond, and proline has none. The older usage that included proline is marked obsolescent; proline is more accurately described as a cyclic secondary amino acid.
Why do proline and glycine break alpha helices?
They do it for opposite reasons. Proline has no backbone N-H to donate a hydrogen bond inside a chain, and its ring fixes the phi angle. Glycine’s effect is usually explained by its flexibility, since fixing it into a helix costs conformational entropy, and model-peptide work found a helix-propensity ratio of about 100 between alanine and glycine.
Why does a proline-containing peptide show two HPLC peaks?
Cis and trans conformers of an X-Pro bond interconvert slowly enough that a reversed-phase column can partly separate them. Studies of X-Pro dipeptides found two peaks at low column temperature that merged into one broad peak as temperature or run time increased. Because both conformers have the same mass, a mass-spectrometry check distinguishes conformer peaks from a genuine impurity.
Proline-rich research peptides, shipped from inside Canada
Semax, Selank and related reagents, priced in Canadian dollars and shipped tracked from inside Canada. Checkout accepts credit and debit cards, Interac e-Transfer and cryptocurrency.
References
- Moss GP, Smith PAS, Tavernier D. Glossary of class names of organic compounds and reactivity intermediates based on structure (IUPAC Recommendations 1995). Pure Appl Chem. 1995;67(8-9):1307–1375. DOI; entry “imino acids”, IUPAC Gold Book
- National Center for Biotechnology Information. PubChem Compound Summary for CID 750, Glycine: dissociation constants. PubChem
- National Center for Biotechnology Information. PubChem Compound Summary for CID 145742, L-Proline: dissociation constants. PubChem
- Matsumoto A, Ozaki H, Tsuchiya S, et al. Achiral amino acid glycine acts as an origin of homochirality in asymmetric autocatalysis. Org Biomol Chem. 2019;17(17):4200–4203. DOI
- Ho BK, Brasseur R. The Ramachandran plots of glycine and pre-proline. BMC Struct Biol. 2005;5:14. DOI
- Chakrabartty A, Schellman JA, Baldwin RL. Large differences in the helix propensities of alanine and glycine. Nature. 1991;351:586–588. DOI
- Brandts JF, Halvorson HR, Brennan M. Consideration of the possibility that the slow step in protein denaturation reactions is due to cis-trans isomerism of proline residues. Biochemistry. 1975;14(22):4953–4963. DOI
- Lang K, Schmid FX, Fischer G. Catalysis of protein folding by prolyl isomerase. Nature. 1987;329:268–270. DOI
- Jabs A, Weiss MS, Hilgenfeld R. Non-proline cis peptide bonds in proteins. J Mol Biol. 1999;286(1):291–304. DOI
- Craveur P, Joseph AP, Poulain P, de Brevern AG, Rebehmed J. Cis-trans isomerization of omega dihedrals in proteins. Amino Acids. 2013;45(2):279–289. DOI
- Alderson TR, Lee JH, Charlier C, Ying J, Bax A. Propensity for cis-proline formation in unfolded proteins. ChemBioChem. 2018;19(1):37–42. DOI
- Nishikawa T, Hayashi Y, Suzuki S, Kubo H, Ohtani H. Cis-trans isomerization of proline dipeptides during liquid chromatography: kinetic analysis of the elution profile. Anal Sci. 1996;12(4):561–564. DOI
- Jenkins CL, Raines RT. Insights on the conformational stability of collagen. Nat Prod Rep. 2002;19(1):49–59. DOI
- Shoulders MD, Raines RT. Collagen structure and stability. Annu Rev Biochem. 2009;78:929–958. DOI
- Stapley BJ, Creamer TP. A survey of left-handed polyproline II helices. Protein Sci. 1999;8(3):587–595. DOI
- Pickart L, Margolina A. Regenerative and protective actions of the GHK-Cu peptide in the light of the new gene data. Int J Mol Sci. 2018;19(7):1987. DOI
For laboratory research use only. Not for human or veterinary use. This article summarizes published research for educational purposes and is not medical advice.
