Henry was born in Studley, Hanover County, Virginia on May 29, 1736. His father was John Henry, an immigrant from Aberdeenshire, Scotland, who had attended King's College, Aberdeen, before immigrating to the Colony of Virginia in the 1720s. Settling in Hanover County, about 1732 John Henry married Sarah Winston Syme, a wealthy widow from a prominent Hanover County family of English ancestry. Patrick Henry was once thought to have been of humble origins, but he was actually born into the middle rank of the Virginia gentry.

Henry attended local schools for a few years, and then was tutored by his father. After failing in business, in 1754 he married Sarah Shelton, with whom he would have six children. As a wedding gift his father-in-law gave the couple six slaves and the 300-acre Pine Slash Farm. Henry began a career as a planter, but their home was destroyed by fire in 1757. Henry made another attempt at business, which also failed, before deciding to become a lawyer in 1760.

Henry first made a name for himself in a case dubbed the "Parson's Cause" (1763), which was an argument about whether the price of tobacco paid to clergy for their services should be set by the colonial government or by the Crown. After the British Parliament overruled Virginia's Two Penny Act that had limited the clergy's salaries, the Reverend James Maury filed suit against the vestry of Louisa County for payment of back wages. When Maury won the suit, a jury was called in Hanover County to determine how much Maury should be paid. Henry was brought in at the last minute to argue on behalf of Louisa County. Ignoring legal niceties, Henry delivered an impassioned speech that denounced clerics who challenged Virginia's laws as "enemies of the community" and any king who annulled good laws like the Two Penny Act as a "tyrant" who "forfeits all right to his subject's obedience". Henry urged the jury to make an example of Maury. After less than five minutes of deliberation, they awarded Maury one penny.

Patrick Henry was elected from Louisa County to the House of Burgesses, the legislative body of the Virginia colony in 1765 to fill a vacated seat in the assembly. When he arrived in Williamsburg the legislature was already in session. Only nine days after being sworn in Henry introduced the Virginia Stamp Act Resolutions, "in language so extreme that some Virginians said it smacked of treason". The freshman representative waited for an opportunity where the mostly conservative members of the House were away (only 24% was considered sufficient for a quorum). In this atmosphere, he succeeded, through much debate and persuasion, in getting his proposal passed. It was possibly the most anti-British American political action to that point, and some credit the Resolutions with being one of the main catalysts of the Revolution. The proposals were based on principles that were well-established British rights, such as the right to be taxed by one's own representatives. They went further, however, to assert that the colonial assemblies had the exclusive right to impose taxes on the colonies and could not assign that right. The imputation of treason is due to his inflammatory words, "Caesar had his Brutus; Charles the First his Cromwell; and George the Third may profit by their example. If this be treason, make the most of it!"

According to biographer Richard Beeman, the legend of this speech grew more dramatic over the years. Henry probably did not say the famous last line of the above quote, i.e. "If this be treason, make the most of it." The only account of the speech written down at the time by an eyewitness (which came to light many years later) records that Henry actually apologized after being accused of uttering treasonable words, assuring the House that he was still loyal to the king. Nevertheless, Henry's passionate, radical speech was cause for notable interest at the time – even if his exact words are unknown.

Patrick Henry is perhaps best known for the speech he made in the House of Burgesses on March 23, 1775, in Saint John's Church in Richmond, Virginia. The House was undecided on whether to mobilize for military action against the encroaching British military force, and Henry argued in favor of mobilization. Forty-two years later, Henry's first biographer, William Wirt, working from oral testimony, attempted to reconstruct what Henry said. According to Wirt, Henry ended his speech with words that have since become immortalized:

Is life so dear, or peace so sweet, as to be purchased at the price of chains and slavery? Forbid it, Almighty God! I know not what course others may take; but as for me, Give me Liberty, or give me Death!

The crowd, by Wirt's account, jumped up and shouted "To Arms! To Arms!". For 160 years Wirt's account was taken at face value, but in the 1970s historians began to question the authenticity of Wirt's reconstruction. Historians today observe that Henry was known to have used fear of Indian and slave revolts in promoting military action against the British, and that according to the only written first-hand account of the speech, Henry used some graphic name-calling that failed to appear in Wirt's heroic rendition.

In August 1775, Henry became colonel of the 1st Virginia Regiment. At the outset of the Revolutionary War, Henry led militia against Royal Governor Lord Dunmore in defense of some disputed gunpowder, an event known as the Gunpowder Incident. During the war he served as the first post-colonial Governor of Virginia and presided over several invasions of Cherokee Indian lands.

Henry lived during part of the War at his 10,000-acre (40 km²) Leatherwood Plantation in Henry County, Virginia, where he, his first cousin Ann Winston Carr and her husband Col. George Waller had settled. During the five years Henry lived at Leatherwood, from 1779 to 1784, he owned 75 slaves, and grew tobacco. During this time, he kept in close touch with his friend the explorer Joseph Martin, whom Henry had appointed agent to the Cherokee nation, and with whom Henry sometimes invested in real estate, and for whom the county seat of Henry County was later named.

In early November 1775 Henry and James Madison were elected founding trustees of Hampden-Sydney College, which opened for classes on November 10. He remained a trustee until his death in 1799. Henry was instrumental in achieving passage of the College's Charter of 1783, an action delayed because of the war. He is probably the author of the Oath of Loyalty to the new Republic included in that charter. Seven of his sons attended the new college.

On October 25, 1777, Patrick Henry married his second wife, Dorothea Dandridge (1755 – 1831). From this marriage came eleven children.

After the Revolution, Henry again served as governor of Virginia from 1784 to 1786, but declined to attend the Constitutional Convention of 1787 saying that he "smelt a rat in Philadelphia, tending toward the monarchy." An ardent supporter of state rights, Henry was an outspoken critic of the United States Constitution and led the Virginia opposition to its ratification arguing that it gave the federal government too much power and that the untested office of the presidency could devolve into a monarchy. As a leading Antifederalist, he was instrumental in forcing the adoption of the Bill of Rights to amend the new Constitution and became a leading opponent of James Madison.

Henry served as a representative to the Virginia convention of 1788 that ratified the U. S. Constitution. He voted against ratification. He was chosen as an elector for the 1789 election from Campbell District. That District consisted of Bedford County, Campbell County, Charlotte County, Franklin County, Halifax County, Henry County, Pittsylvania County, and Prince Edward County, which cover the area between Danville and Lynchburg in the south of Virginia. All of the 10 electors who voted cast one of their two votes for George Washington. Five of them cast their other vote for John Adams. Three cast theirs for George Clinton. One cast his for John Hancock. One cast his for John Jay. Clinton was a leading Antifederalist, a view which he shared with Henry.

President George Washington offered Henry the post of Secretary of State in 1795, which he declined out of opposition to Washington's Federalist policies. However, following the radicalism of the French Revolution Henry's views changed as he began to fear a similar fate could befall America and by the late 1790s Henry was in support of the Federalist policies of Washington and Adams. He especially denounced the Virginia and Kentucky Resolutions, which had been secretly written by Jefferson and Madison, and approved by the legislatures of those two states. He warned that civil war was threatened because Virginia, "had quitted the sphere in which she had been placed by the Constitution, and, in daring to pronounce upon the validity of federal laws, had gone out of her jurisdiction in a manner not warranted by any authority, and in the highest degree alarming to every considerate man; that such opposition, on the part of Virginia, to the acts of the general government, must beget their enforcement by military power; that this would probably produce civil war, civil war foreign alliances, and that foreign alliances must necessarily end in subjugation to the powers called in." In 1798 President John Adams nominated Henry special emissary to France, but he had to decline because of failing health. He strongly supported John Marshall and at the urging of Washington stood for and was elected to the Virginia House of Delegates as a Federalist. However, three months prior to taking his seat in the state legislature, he died of stomach cancer on June 6, 1799, while at Red Hill, his family's large plantation. Following his death, Henry's widow remarried the Virginia governor's first cousin and executor Judge Edmund Winston.